Gene (v.498, #2)
Regulation of the human catalytic subunit of telomerase (hTERT)
by Michael Daniel; Gregory W. Peek; Trygve O. Tollefsbol (pp. 135-146).
Over the past decade, there has been much interest in the regulation of telomerase, the enzyme responsible for maintaining the integrity of chromosomal ends, and its crucial role in cellular immortalization, tumorigenesis, and the progression of cancer. Telomerase activity is characterized by the expression of the telomerase reverse transcriptase ( TERT) gene, suggesting that TERT serves as the major limiting agent for telomerase activity. Recent discoveries have led to characterization of various interactants that aid in the regulation of human TERT ( hTERT), including numerous transcription factors; further supporting the pivotal role that transcription plays in both the expression and repression of telomerase. Several studies have suggested that epigenetic modulation of the hTERT core promoter region may provide an additional level of regulation. Although these studies have provided essential information on the regulation of hTERT, there has been ambiguity of the role of methylation within the core promoter region and the subsequent binding of various activating and repressive agents. As a result, we found it necessary to consolidate and summarize these recent developments and elucidate these discrepancies. In this review, we focus on the co-regulation of hTERT via transcriptional regulation, the presence or absence of various activators and repressors, as well as the epigenetic pathways of DNA methylation and histone modifications.► Telomerase is regulated by the hTERT gene. ► The hTERT gene is controlled by genetic and epigenetic factors. ► hTERT gene control is important in cancer and aging.
Keywords: Abbreviations; AP1; activator protein 1; AP2; activating protein 2; AP-2β; activating enhancer binding protein-2β; AP4; activating enhancer-binding protein 4; APPB; amyloid beta (A4) precursor protein B; BRCA1; breast cancer type 1 susceptibility protein; CDC25A; cell division cycle 25 homolog A; Cdk; cyclin dependent kinase; CREB; cAMP responsive element binding protein; CTCF; CCCTC-binding factor; DNMT; DNA methyl transferase; E2; estradiol; EGF; epidermal growth factor; EGFR; epidermal growth factor receptor; ERα; estrogen receptor alpha; ERβ; estrogen receptor beta; ErbB; human epidermal growth factor 2 (Her2/neu); ERE; estrogen-response element; ERK; extracellular signal-regulated kinase; HAT; Histone acetylases; HCC; heptocellular carcinoma; HDAC; histone deacetylase; HER2/neu; human epidermal growth factor 2; HMGA2; high-mobility group A2; hTERT; human telomerase reverse transcriptase; IFN-β; Interferon-β; IGF2; insulin-like growth factor 2; IK2; IkappaB kinase-like 2; IKB (I kappaB); inhibitor of kappa B; IKK; inhibitor of kappaB kinase; INHBC; inhibin beta C chain; IRF1; Interferon regulatory factor 1; Jab1; c-Jun activation domain-binding protein-1; MAD1; mitotic spindle assembly checkpoint protein MAD1; MAD1; mitotic spindle assembly checkpoint protein MAD1; MAPK; mitogen-activated protein kinase; MAZ; Myc-associated zinc finger protein; MCAF1–MBD1; containing chromatin-associated factor 1; MDM2; murine double minute 2; MKRN1; Makorin-1; MNT; MAX binding protein; mTOR; mammalian target of rapamycin; MyoD; myoblast determination protein 1; MZF-2; myeloid zinc finger protein 2; NF1; nuclear factor 1; NFAT; nuclear factor of activated T-cells; NF-E2; nuclear factor (erythroid-derived 2); NF-κB; nuclear factor-kappaB; NIK; NF-kappaB-inducing kinase; PIP2; phosphatidylinositol (3,4)-bisphosphate; PIP3; phosphatidylinositol (3,4,5)-trisphosphate; PITX1; paired-like homeodomain 1; PTEN; phosphatase and tensin homolog; Rb; retinoblastoma protein; RSK; ribosomal protein S6 kinase; S6K; S6 kinase; Sp1; specificity protein 1; SMYD3 – SET-and MYND; domain-containing protein-3; TERT; telomerase reverse transcriptase; TEP1; telomerase protein component 1; TFIID; transcription factor IID; TGF-β; transforming growth factor beta; TRF1; telomeric repeat binding factor 1; TRF2; telomeric repeat binding factor 2; TSA; Tricostatin A; USF1; upstream stimulatory factor 1; WTI; Wilms tumor proteinTelomerase; Cancer; Gene regulation; Telomeres
Cloning, localization and differential expression of the Trypanosoma cruzi TcOGNT-2 glycosyl transferase
by María Laura Chiribao; María Gabriela Libisch; Eduardo Osinaga; Adriana Parodi-Talice; Carlos Robello (pp. 147-154).
The surface of Trypanosoma cruzi is covered by a dense glycocalix which is characteristic of each stage of the life cycle. Its composition and complexity depend mainly on mucin-like proteins. A remarkable feature of O-glycan biosynthesis in trypanosomes is that it initiates with the addition of a GlcNAc instead of the GalNAc residue that is commonly used in vertebrate mucins. The fact that the interplay between trans-sialidase and mucin is crucial for pathogenesis, and both families have stage-specific members is also remarkable. Recently the enzyme that transfers the first GlcNAc from UDP-GlcNAc to a serine or threonine residue was kinetically characterized. The relevance of this enzyme is evidenced by its role as catalyzer of the first step in O-glycosylation. In this paper we describe how this gene is expressed differentially along the life cycle with a pattern that is very similar to that of trans-sialidases. Its localization was determined, showing that the protein predicted to be in the Golgi apparatus is also present in reservosomes. Finally our results indicate that this enzyme, when overexpressed, enhances T. cruzi infectivity.► TcOGNT-2 product gene is localized in the Golgi apparatus and reservosomes. ► It is differentially expressed. ► TcOGNT-2 gene is regulated in a similar way that trans-sialidase gene. ► Its overexpression enhance parasite infectivity.
Keywords: Abbreviations; GlcNAc; N-acetylglucosamine; Tc; OGNT-2; Trypanosoma cruzi O; -GlcNAc-transferase 2; GPI; glycosylphosphatidylinositol; Tc; MUC; Trypanosoma cruzi; mucin; TS; trans; -sialidase; DAPI; 4′,6-diamidino-2-phenylindoleChagas disease; Glycosyl-transferase; Differential expression; O; -glycosylation; Infectivity
Characterization of an Abc1 kinase family gene OsABC1- 2 conferring enhanced tolerance to dark-induced stress in rice
by Qingsong Gao; Zefeng Yang; Yong Zhou; Zhitong Yin; Jie Qiu; Guohua Liang; Chenwu Xu (pp. 155-163).
Leaf senescence is a complex and highly organized process resulting in numerous changes of gene expression and metabolic procedures. However, the exact mechanisms underlying these changes are not well understood. In this study, we reported a rice ( Oryza sativa) T-DNA insertion mutant impaired in an Abc1 kinase family gene with a dwarf and pale-green phenotype. The mutant showed reduced pigment content and photosynthetic efficiency and increased superoxide dismutase activity in leaves. The mutated gene, designated OsABC1- 2, is expressed primarily in green tissues and/or organs and encodes a protein localized in chloroplast envelope. Expression of the gene was drastically suppressed by dark treatment. Overexpression of the gene in rice enhanced tolerance to prolonged dark-induced stress. Phylogenetic analysis revealed that the plant Abc1 proteins could be divided into three subgroups and OsAbc1-2 resides in a subgroup with potential chloroplast origin. Our results suggest that divergence has occurred among plant Abc1 family and chloroplast Abc1 kinases play potential roles in regulating dark-induced senescence of plants.► A novel Abc1 kinase of Oryza sativa was characterized. ► The OsAbc1-2 protein is localized to chloroplast envelope. ► Overexpression of OsABC1- 2 confers tolerance to prolonged dark-induced stress. ► Chloroplast Abc1 proteins play potential roles in plant adaption to dark stress.
Keywords: Abbreviations; ABA; abscisic acid; Abc1; Activity of bc1 complex; DIG; digoxigenin; GUS; β-; glucuronidase; GFP; green fluorescent protein; MS; Murashige and Skoog; OE; overexpression; ROS; reactive oxygen species; SOD; superoxide dismutase; VE; vitamin E; WT; wild type; ZH11; Zhonghua 11 japonica riceAbc1 kinase; Dark tolerance; Chloroplast; Oryza sativa
Characterization of a Tigger1 element from the genome of the American mink ( Neovison vison)
by Amanda Smith; Katherine Rutherford; Bernhard Benkel (pp. 164-168).
Tigger elements belong to the Tc1/Mariner family of DNA transposons which is remarkably widespread in nature with homologs present in organisms as diverse as fungi, plants and animals. In this report, we present the nucleotide sequence of a defragmented Tigger1 element from the genome of the American mink. The element is 2274bp long, carries 13bp terminal inverted repeats (TIRs) and contains the vestiges of two open reading frames (ORFs), one of which is similar to the centromere associated protein CENP B. In addition, we estimate that the genome of the American mink contains approximately 1000 Tigger1 elements, but find no evidence for the transcription of extant elements in the mink.► 1000 Tigger1 elements in mink genome. ► No evidence for expression. ► Ancient resident of mink genome.
Keywords: Abbreviations; α-; 32; P; alpha Phosphorus 32; BAC; bacterial artificial chromosome; bp; base pair; BES; BAC end sequence; Ci; curies; °C; degree Celcius; dCTP; deoxycytidine triphosphate; DNA; deoxyribonucleic acid; dNTP; deoxyribonucleoside triphosphate; Gb; gigabase; hr; hour; LR-IPCR; long range inverse PCR; LR-PCR; long range PCR; M; molar; ug; microgram; min; minute; mg; milligram; mL; milliliter; mM; millimolar; Mb; megabase; mya; million years ago; N; normal; ng; nanogram; nm; nanometers; nM; nanomolar; ORF; open reading frame; PCR; polymerase chain reaction; polyA; +; RNA; polyadenylated RNA; RNA; ribonucleic acid; RT; room temperature; s; seconds; SDS; sodium dodecyl sulfate; SINE; short interspersed nuclear element; SSC; 0.15; M NaCl/0.015; M Na; 3; citrate pH 7.6; TE; transposable element; TIR; terminal inverted repeat; U; units; UV; ultraviolet light; v/v; volume per volumeDNA transposons; Mustelids
Three novel BRCA1/ BRCA2 mutations in breast/ovarian cancer families in Croatia
by Sonja Levanat; Vesna Musani; Mirela Levacic Cvok; Ilona Susac; Maja Sabol; Petar Ozretic; Diana Car; Domagoj Eljuga; Ljerka Eljuga; Damir Eljuga (pp. 169-176).
BRCA1 and BRCA2 genes from 167 candidates (145 families) were scanned for mutations. We identified 14 pathogenic point mutations in 17 candidates, 9 in BRCA1 and 5 in BRCA2. Of those, 11 have been previously described and 3 were novel (c.5335C>T in BRCA1 and c.4139_4140dupTT and c.8175G>A in BRCA2). No large deletions or duplications involving BRCA1 and BRCA2 genes were identified. No founder mutations were detected for the Croatian population. Croatia shares most of the mutations with neighboring Slovenia and also with Germany, Austria and Poland.Two common sequence variants in BRCA1, c.2077G>A and c.4956G>A, were found more frequently in mutation carriers compared to healthy controls. No difference in BRCA2 variants was detected between the groups.Haplotype inference showed no difference in haplotype distributions between deleterious mutation carriers and non-carriers in neither BRCA1 nor BRCA2. In silico analyses identified one BRCA1 sequence variant (c.4039A>G) and two BRCA2 variants (c.5986G>A and c.6884G>C) as harmful with high probability, and inconclusive results were obtained for our novel BRCA2 variant c.3864_3866delTAA.Combination of QMPSF and HRMA methods provides high detection rate and complete coverage of BRCA1/2 genes. Benefit of BRCA1/2 mutation testing is clear, since we detected mutations in young unaffected women, who will be closely monitored for breast and ovarian cancer.► QMPSF and HRMA provide high detection rate and complete coverage of BRCA1/2 genes. ► 14 pathogenic point mutations in 17 candidates, 9 in BRCA1 and 5 in BRCA2. ► Three novel mutations: BRCA1 c.5335C>T; BRCA2 c.4139_4140dupTT and c.8175G>A. ► BRCA1 c.2077G>A and c.4956G>A variants are more frequent among mutation carriers. ► In silico analysis confirmed BRCA2 mutation c.9371A>T pathogenic character.
Keywords: BRCA1; BRCA2; hereditary breast/ovarian cancer; high-resolution melting analysis; quantitative multiplex PCR of short fluorescent fragments
Functional study of the novel multidrug efflux pump KexD from Klebsiella pneumoniae
by Wakano Ogawa; Motoyasu Onishi; Ruiting Ni; Tomofusa Tsuchiya; Teruo Kuroda (pp. 177-182).
We cloned a gene, kexD, that provides a multidrug-resistant phenotype from multidrug-resistant Klebsiella pneumoniae MGH78578. The deduced amino acid sequence of KexD is similar to that of the inner membrane protein, RND-type multidrug efflux pump. Introduction of the kexD gene into Escherichia coli KAM32 resulted in a MIC that was higher for erythromycin, novobiocin, rhodamine 6G, tetraphenylphosphonium chloride, and ethidium bromide than that of the control. Intracellular ethidium bromide levels in E. coli cells carrying the kexD gene were lower than that in the control cells under energized conditions, suggesting that KexD is a component of an energy-dependent efflux pump. RND-type pumps typically consist of three components: an inner membrane protein, a periplasmic protein, and an outer membrane protein. We discovered that KexD functions with a periplasmic protein, AcrA, from E. coli and K. pneumoniae, but not with the periplasmic proteins KexA and KexG from K. pneumoniae. KexD was able to utilize either TolC of E. coli or KocC of K. pneumoniae as an outer membrane component. kexD mRNA was not detected in K. pneumoniae MGH78578 or ATCC10031. We isolated erythromycin-resistant mutants from K. pneumoniae ATCC10031, and some showed a multidrug-resistant phenotype similar to the drug resistance pattern of KexD. Two strains of multidrug-resistant mutants were investigated for kexD expression; kexD mRNA levels were increased in these strains. We conclude that changing kexD expression can contribute to the occurrence of multidrug-resistant K. pneumoniae.► We characterized an RND-type multidrug efflux pump, KexD, from K. pneumoniae. ► KexD can function with either AcrA Ec or AcrA Kp, which are periplasmic proteins. ► KexD also requires either TolC from E. coli or KocC from K. pneumoniae. ► Drug-resistant mutants with increased kexD expression were isolated . ► We conclude that kexD contributes to acquired resistance in K. pneumoniae.
Keywords: Abbreviations; SDS; sodium dodecyl sulfate; TPPCl; tetraphenylphosphonium; Hoechst 33342; 2′-[4-ethoxyphenyl]-5-[4-methyl-1-piperazinyl]-2,5′-bis-1H-benzimidazole; EtBr; ethidium bromide; NFLX; norfloxacin; CCCP; carbonyl cyanide m-chlorophenylhydrazone; CTAB; cetyl trimethyl ammonium bromide; RND; resistance; −; nodulation; −; cell division; SMR; small multidrug resistance; MATE; multidrug and toxic compounds extrusion; MF; major facilitator; ABC; ATP binding cassette; MIC; minimum inhibitory concentration; PCR; polymerase chain reaction; AcrA; Kp; AcrA from; K. pneumoniae; AcrA; Ec; AcrA from; E. coliRND-type efflux pump; Multidrug resistance; Klebsiella pneumoniae
Danon disease: A focus on processing of the novel LAMP2 mutation and comments on the beneficial use of peripheral white blood cells in the diagnosis of LAMP2 deficiency
by F. Majer; H. Vlaskova; L. Krol; T. Kalina; M. Kubanek; L. Stolnaya; L. Dvorakova; M. Elleder; J. Sikora (pp. 183-195).
Danon disease (DD) is a monogenic X-linked disorder characterized by cardiomyopathy, skeletal myopathy and variable degrees of intellectual disability. DD develops due to mutations in the gene encoding lysosomal-associated membrane protein 2 (LAMP2). We report on a family exhibiting the clinical phenotype comprising of hypertrophic cardiomyopathy and ventricular pre-excitation, myopia and mild myopathy in two male patients and cardiomyopathy and myopia in a female patient. The diagnosis of DD in this family was based on the assessment of the clinical phenotypes and the absence of LAMP2 in skeletal and/or cardiac muscle biopsy specimens. Sequence analysis of the LAMP2 gene and its mRNA revealed a novel LAMP2 mutation ( c.940delG) in all three patients.Approximately 25% of the female patient's cardiomyocytes were LAMP2 positive apparently due to the unfavorable skewing of X chromosome inactivation. We further performed qualitative LAMP2 immunohistochemistry on peripheral white blood cells using the smear technique and revealed the absence of LAMP2 in the male patients. LAMP2 expression was further assessed in granulocytes, CD4+ and CD8+ T lymphocytes, CD20+ B lymphocytes, CD14+ monocytes and CD56+ natural killer cells by quantitative polychromatic flow cytometry. Whereas the male DD patients lacked LAMP2 in all WBC populations, the female patient expressed LAMP2 in 15.1% and 12.8% of monocytes and granulocytes, respectively. LAMP2 expression ratiometrics of highly vs. weakly expressing WBC populations discriminated the DD patients from the healthy controls. WBCs are thus suitable for initial LAMP2 expression testing when DD is a differential diagnostic option. Moreover, flow cytometry represents a quantitative method to assess the skewing of LAMP2 expression in female heterozygotes. Because LAMP2 is a major protein constituent of the membranes of a number of lysosome-related organelles, we also tested the exocytic capacity of the lytic granules from CD8+ T lymphocytes in the patient samples. The degranulation triggered by a specific stimulus (anti-CD3 antibody) was normal. Therefore, this process can be considered LAMP2 independent in human T cells.The c.940delG mutation results in a putatively truncated protein (p.A314QfsX32), which lacks the transmembrane domain and the cytosolic tail of the wild-type LAMP2. We tested whether this variant becomes exocytosed because of a failure in targeting to late endosomes/lysosomes. Western blotting of cardiac muscle, WBCs and cultured skin fibroblasts (and their culture media) showed no intra- or extracellular truncated LAMP2. By comparing the expression pattern and intracellular targeting in cultured skin fibroblasts of normal LAMP2 isoforms (A, B and C) tagged with green fluorescent protein (GFP) and the A314Qfs32-GFP fusion, we found that the A314Qfs32-GFP protein is not even expressed. These observations suggest that the truncated protein is unstable and is co-translationally or early post-translationally degraded.► c.940delG (p.A314QfsX32) is a novel Danon disease associated LAMP2 gene mutation. ► A314Qfs32 LAMP2 variant is degraded and is not secreted from cells. ► White blood cells should be preferably tested for LAMP2 expression in Danon disease. ► Flow cytometry allows assessment of skewed expression of LAMP2 ( Xq24) alleles. ► Exocytosis of lytic granules is LAMP2 independent in human CD8+ T lymphocytes.
Keywords: Abbreviations; aa; amino acid; Ab; antibody; AP; autophagosomes; CMA; chaperone-mediated autophagy; DD; Danon disease; FFPE; formalin-fixed paraffin embedded; FMO; Fluorescence minus one; GFP; green fluorescent protein; HCM; hypertrophic cardiomyopathy; LAMP2; lysosomal-associated membrane protein 2; LRO; lysosome-related organelles; LV; left ventricle; PhG; phagosomes; RV; right ventricle; TM; transmembrane; WB; Western blotting; WBC; white blood cells; XCI; X-chromosome inactivationDanon disease; Lysosomal-associated membrane protein 2; Cardiomyopathy; White blood cells; Polychromatic flow cytometry; X-chromosome inactivation
Molecular structure, expression analysis and functional characterization of APRIL (TNFSF13) gene in bat ( Vespertilio superans Thomas)
by Fengtao You; Lidan Zhou; Xuanxuan Liu; Jie Fan; Zhen Ke; Wenhua Ren (pp. 196-202).
A proliferation-inducing ligand (APRIL) is a novel member of the tumor necrosis factor (TNF) superfamily, which is involved in immune regulation. In the present study, the full-length cDNA of APRIL (designated bAPRIL) from bat was cloned using RT-PCR and its biological activities have been characterized. The open reading frame (ORF) of this cDNA consists of 753 bases, encoding a protein of 250 amino acids. This protein was found to contain a predicted transmembrane domain, a putative furin protease cleavage site, and a typical TNF homology domain corresponding to other, known APRIL homologs. Real-time quantitative PCR (qPCR) analysis indicated that bAPRIL mRNA was predominantly expressed in bat lymphoid tissue spleen. The SUMO–bsAPRIL was efficiently expressed in Escherichia coli BL21 (DE3) and confirmed by SDS-PAGE and Western blot analysis. Laser scanning confocal microscopy analysis showed that bsAPRIL could bind to its receptors on B cells. In vitro, MTT assays indicated that bsAPRIL could promote the survival/proliferation of mouse splenic B cells grown with anti-mouse IgM. These findings indicate that bsAPRIL plays an important role in the survival and proliferation of B cells and has functional cross-reactivity among mammalians. The present findings may provide valuable information for research into the immune system of the bat.► We successfully cloned the APRIL gene in bat ( Vespertilio superans Thomas). ► The deduced amino acid sequence was analyzed using bioinformatics methods. ► Real-time quantitative PCR indicated APRIL mRNA was predominantly expressed in spleen. ► Protein was expressed in E. coli and purified by Ni column. ► MTT analysis tested the bioactivity of APRIL to B cells.
Keywords: Abbreviations; APRIL; A proliferation-inducing ligand; bp; base pair(s); BAFF; B cell activating factor; BCMA; B cell maturation antigen; cDNA; DNA complementary; DMSO; dimethylsulfoxide; dNTP; deoxyribonucleoside triphosphate; HSPG; heparin sulfate proteoglycan; IPTG; final isopropyl-beta-; d; -thiogalactopyranoside; kDa; kilodalton(s); LB; Luria-Bertani (medium); FBS; fetal bovine serum; FITC; fluoresceine isothiocyanate; MTT; 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; ORF; open reading frame; PBS; phosphate buffered saline; PCR; polymerase chain reaction; PI; isoelectric point; qPCR; real-time quantitative PCR; SDS-PAGE; sodium dodecyl sulfate-polyacrylamide electrophoresis; SUMO; small ubiquitin-like modifier; TACI; transmembrane activator and cyclophilin ligand interactor; TMD; transmembrane domain; TNF; tumor necrosis factorAPRIL; Bat (; Vespertilio superans; Thomas); Phylogenetic analysis; MTT assay; B cell survival
ESTs library from embryonic stages reveals tubulin and reflectin diversity in Sepia officinalis (Mollusca — Cephalopoda)
by Yann Bassaglia; Thomas Bekel; Corinne Da Silva; Julie Poulain; Aude Andouche; Sandra Navet; Laure Bonnaud (pp. 203-211).
New molecular resources regarding the so-called “non-standard models” in biology extend the present knowledge and are essential for molecular evolution and diversity studies (especially during the development) and evolutionary inferences about these zoological groups, or more practically for their fruitful management. Sepia officinalis, an economically important cephalopod species, is emerging as a new lophotrochozoan developmental model. We developed a large set of expressed sequence tags (ESTs) from embryonic stages of S. officinalis, yielding 19,780 non-redundant sequences (NRS). Around 75% of these sequences have no homologs in existing available databases. This set is the first developmental ESTs library in cephalopods. By exploring these NRS for tubulin, a generic protein family, and reflectin, a cephalopod specific protein family, we point out for both families a striking molecular diversity in S. officinalis.► The first ESTs library from developmental stages of a cephalopod species is described. ► 75% of the 19,780 unigenes of Sepia officinalis have no homology in databases. ► The tubulin family is highly diverse and diverged into the cephalopod clade. ► The reflectin repertoire covers all reflectin forms known in other cephalopod species.
Keywords: Abbreviation; ESTs; expressed sequence tags; NRS; non-redundant sequences; TCs; tentative contigs; KOG; eukaryotic orthologous groups; SAMS; Sequence Analysis and Management System; JGI; Joint Genome Institute; aa; amino acid Sepia officinalis; Development; Expressed sequence tags (ESTs); Tubulin; Reflectin
Genome-wide analysis and environmental response profiling of the FK506-binding protein gene family in maize ( Zea mays L.)
by Yanli Yu; Hui Zhang; Wencai Li; Chunhua Mu; Fajun Zhang; Liming Wang; Zhaodong Meng (pp. 212-222).
The FK506-binding proteins (FKBPs) belong to the peptidyl-prolyl cis/trans isomerase (PPIase) superfamily, and have been implicated in a wide spectrum of biological processes, including protein folding, hormone signaling, plant growth, and stress responses. Genome-wide structural and evolutionary analyses of the entire FKBP gene family have been conducted in Arabidopsis and rice. In the present study, a genome-wide analysis was performed to identify all maize FKBP genes. The availability of complete maize genome sequences allowed for the identification of 24 FKBP genes. Chromosomal locations in the maize genome were determined and the protein domain and motif organization of ZmFKBPs analyzed. The phylogenetic relationships between maize FKBPs were also assessed. The expression profiles of ZmFKBP genes were measured under different environmental conditions and revealed distinct ZmFKBP gene expression patterns under heat, cold, salt, and drought stress. These data not only contribute to a better understanding of the complex regulation of the maize FKBP gene family, but also provide evidence supporting the role of FKBPs in multiple signaling pathways involved in stress responses. This investigation may provide valuable information for further research on stress tolerance in plants and potential strategies for enhancing maize survival under stressful conditions.► FKBP genes in maize were identified for the first time. ► An overview of FKBP gene family in maize was presented. ► The expression profiles of ZmFKBP genes were presented. ► Several ZmFKBPs exhibit different expression levels in various stress conditions.
Keywords: Abbreviations; FKBPs; FK506-binding proteins; FKBd; FK506-binding domain; PPIase; peptidyl-prolyl cis/trans isomerase; TPR; tetratricopeptide repeat; CCD; coiled-coil domain; CaM-BDs; calmodulin-binding domains; SD; single-domain; MD; multidomain; TOR; target of rapamycin; HSP90; heat streak protein 90; CDS; coding sequence; PlantGBD; Plant Genomic Database; PUTs; PlantGDB unique transcripts; RT-PCR; semi-quantitative reverse transcription PCR; qRT-PCR; quantitative real-time PCR; ER; endoplasmic reticulum; ABC; ATP-binding cassetteMaize; FKBPs; Abiotic stress; Signaling pathway
The transcriptomes of columnar and standard type apple trees ( Malus x domestica) — A comparative study
by Clemens Krost; Romina Petersen; Erwin R. Schmidt (pp. 223-230).
Columnar apple trees ( Malus x domestica) provide several economic advantages due to their specific growth habit. The columnar phenotype is the result of the dominant allele of the gene Co and is characterized by thick stems with short internodes and reduced lateral branching. Co is located on chromosome 10 and often appears in a heterozygous state ( Co/ co). The molecular explanation of columnar growth is not well established. Therefore, we studied the transcriptomes of columnar and standard type apple trees using 454 and Illumina next generation sequencing (NGS) technologies. We analyzed the transcriptomes of shoot apical meristems (SAMs) because we expect that these organs are involved in forming the columnar growth phenotype. The results of the comparative transcriptome analysis show significant differences in expression levels of hundreds of genes. Many of the differentially expressed genes are associated with membrane and cell wall growth or modification and can be brought in line with the columnar phenotype. Additionally, earlier findings on the hormonal state of shoots of columnar apples could be affirmed. Our study resulted in a large number of genes differentially expressed in columnar vs. standard type apple tree SAMs. Although we have not unraveled the nature of the Co gene, we could show that the modified expression of these genes, most likely due to the presence of Co, can determine the columnar phenotype. Furthermore, the usefulness of NGS for the analysis of the molecular basis of complex phenotypes is discussed.►The transcriptome of the apical meristems of apple tree was analyzed by NGS. ►Genes differentially expressed in mutant (columnar) apple tree were identified. ►Genes for membrane and cell wall growth are up regulated in columnar tree meristems. ►Up/down regulated genes correlate to the hormonal state typical for columnar trees.
Keywords: Abbreviation; Co; Columnar; NGS; next generation sequencing; SAM; shoot apical meristem; ABA; abscisic acid; IAA; indoleacetic acid; GA; gibberellic acid; A14; ‘A14-190-93’ (standard type apple); P28; ‘Procats28’ (columnar type apple); BORT; Blast Output Refinement Tool; JA; jasmonate Co; gene; NGS; Illumina; 454; De novo assembly; MAPMAN
Lentivirus vector driven by polybiquitin C promoter without woodchuck posttranscriptional regulatory element and central polypurine tract generates low level and short-lived reporter gene expression
by Siew Ching Ngai; Rozita Rosli; Norshariza Nordin; Abhi Veerakumarasivam; Syahril Abdullah (pp. 231-236).
Lentivirus (LV) encoding woodchuck posttranscriptional regulatory element (WPRE) and central polypurine tract (cPPT) driven by CMV promoter have been proven to act synergistically to increase both transduction efficiency and gene expression. However, the inclusion of WPRE and cPPT in a lentiviral construct may pose safety risks when administered to human. A simple lentiviral construct driven by an alternative promoter with proven extended duration of gene expression without the two regulatory elements would be free from the risks. In a non-viral gene delivery context, gene expression driven by human polybiquitin C (UbC) promoter resulted in higher and more persistent expression in mouse as compared to cytomegalovirus (CMV) promoter. In this study, we measured the efficiency and persistency of green fluorescent protein (GFP) reporter gene expression in cells transduced with LV driven by UbC (LV/UbC/GFP) devoid of the WPRE and cPPT in comparison to the established LV construct encoding WPRE and cPPT, driven by CMV promoter (LV/CMV/GFP). However, we found that LV/UbC/GFP was inferior to LV/CMV/GFP in many aspects: (i) the titer of virus produced; (ii) the levels of reporter gene expression when MOI value was standardized; and (iii) the transduction efficiency in different cell types. The duration of reporter gene expression in selected cell lines was also determined. While the GFP expression in cells transduced with LV/CMV/GFP persisted throughout the experimental period of 14days, expression in cells transduced with LV/UbC/GFP declined by day 2 post-transduction. In summary, the LV driven by the UbC promoter without the WPRE and cPPT does not exhibit enhanced or durable transgene expression.► We compared gene expression from two lentiviruses with distinct regulatory elements. ► Lentivirus with CMV, WPRE and cPPT exhibits prolonged and high gene expression. ► Gene expression from lentivirus with UbC without WPRE and cPPT was inferior. ► WPRE and cPPT are necessary for prolonged and high gene expression.
Keywords: Abbreviations; ATCC; American Type Culture Collection; CMV; cytomegalovirus; cPPT; central polypurine tract; DMEM; Dulbecco's modified Eagle's medium; FBS; fetal bovine serum; FIV; feline immunodeficiency virus; GFP; green fluorescent protein; GM-CSF; granulocyte macrophages colony stimulating factor; GMEM; Glasgow minimum essential medium; HIV-1; human immunodeficiency virus type-1; LIF; leukemia inhibitory factor; LV; lentiviral vector; MOI; multiplicity of infection; PBS; phosphate buffer saline; RV; retroviral vector; UbC; polybiquitin C; VSV-G; vesicular stomatitis virus G glycoprotein; WHV; woodchuck hepatitis virus; WPRE; woodchuck posttranscriptional regulatory elementLentivirus; Human ubiquitin C promoter; Green fluorescent protein; Cytomegalovirus promoter; Woodchuck posttranscriptional regulatory element; Central polypurine tract
Prenatal diagnosis of partial trisomy 3q resulting from t(3;14) in a fetus with multiple anomalies including vermian hypoplasia
by Sang Hee Jung; Sung Han Shim; Sang Hee Park; Ji Eun Park; Hea Ree Park; Eun Hee Ahn; Soo Hyun Kim; Dong Hyun Cha (pp. 237-241).
While genetic origin of Dandy–Walker complex has not yet fully elucidated, the complex has been known to be associated with structural and chromosomal abnormalities. A partial trisomy 3q was also identified in patients with DWC.3q duplication syndrome is defined as duplications of large parts of 3q, especially 3q21-qter. Most cases with 3q duplication are diagnosed postnatally and the patients show typical features including various facial dysmorphisms, congenital heart defects, genitourinary malformations, and mental and growth retardation.Here we report a 28year old nulliparous woman who was referred from the infertility clinic at 21 gestational weeks. Fetal ultrasonographic examination showed various abnormal findings including a ventricular septal defect, hydrocephalus, and hypoplasia of the cerebellar vermis. Fetal chromosome analysis was initially reported as 46,XY,der(14)(?::p11.2→qter). Array CGH followed by FISH allowed precise characterization of the der(14) chromosome and the initial karyotype of the fetus had been changed to 46,XY,add(14)(p11).ish der(14)t(3;14)(q26.1;p11)(tel3q+).arr 3q26.1q29(166249469–199288361)x3.Though further studies are required, gene clusters rather than a single gene might be responsible for the clinical features of the Dandy–Walker complex.► Dandy Walker complex (DWC) might be associated with structural and chromosomal abnormalities. ► A partial trisomy 3q was also identified in patients with DWC. ► A trisomy 3q was prenatally diagnosed by array CGH.
Keywords: Abbreviations; DWC; dandy walker complex; DWM; dandy walker malformation; MRI; magnetic resonance imaging; CGH; comparative genomic hybridization; FISH; fluorescence; in situ; hybridization; OMIM; online Mendelian inheritance in man databaseDandy–Walker complex; Partial trisomy 3q; 3q duplication syndrome
A complex multilevel attack on Pseudomonas aeruginosa algT/ U expression and AlgT/U activity results in the loss of alginate production
by Robert Sautter; Damaris Ramos; Lisa Schneper; Oana Ciofu; Tina Wassermann; Chong-Lek Koh; Arne Heydorn; Morton Hentzer; Hoiby Niels Høiby; Arsalan Kharazmi; Søren Molin; Caroline A. DeVries; Dennis E. Ohman; Kalai Mathee (pp. 242-253).
Infection by the opportunistic pathogen Pseudomonas aeruginosa is a leading cause of morbidity and mortality seen in cystic fibrosis (CF) patients. This is mainly due to the genotypic and phenotypic changes of the bacteria that cause conversion from a typical nonmucoid to a mucoid form in the CF lung. Mucoid conversion is indicative of overproduction of a capsule-like polysaccharide called alginate. The alginate-overproducing (Alg+) mucoid phenotype seen in the CF isolates is extremely unstable. Low oxygen tension growth of mucoid variants readily selects for nonmucoid variants. The switching off mechanism has been mapped to the algT/ U locus, and the molecular basis for this conversion was partially attributed to mutations in the algT/ U gene itself. To further characterize molecular changes resulting in the unstable phenotype, an isogenic PAO1 derivative that is constitutively Alg+ due to the replacement of the mucA with mucA22 (PDO300) was used. The mucA22 allele is common in mucoid CF isolates. Thirty-four spontaneous nonmucoid variants, or sap ( suppressor of alginate production) mutants, of PDO300 were isolated under low oxygen tension. About 40% of the sap mutants were rescued by a plasmid carrying algT/ U (Group A). The remaining sap mutants were not (Group B). The members of Group B fall into two subsets: one similar to PAO1, and another comparable to PDO300. Sequence analysis of the algT/ U and mucA genes in Group A shows that mucA22 is intact, whereas algT/ U contains mutations. Genetic complementation and sequencing of one Group B sap mutant, sap22, revealed that the nonmucoid phenotype was due to the presence of a mutation in PA3257. PA3257 encodes a putative periplasmic protease. Mutation of PA3257 resulted in decreased algT/ U expression. Thus, inhibition of algT/ U is a primary mechanism for alginate synthesis suppression.► We screened for genes involved in alginate production. ► Multiple pathways are involved in mucoid to nonmucoid reversion. ► Reversion is in part attributed to loss of AlgT/U and AlgO function. ► AlgT/U and AlgO positively regulate alginate production.
Keywords: Abbreviations; A; adenosine; Ala; alanine; Alg; −; nonmucoid; Alg; +; alginate overproducing, mucoid; ANOLEA; atomic non-local environment assessment; Ap; ampicillin; Asp; aspartic acid; bp; base pair; C; cytidine; Cb; carbenicillin; CF; cystic fibrosis; DNA; deoxyribonucleic acid; F; phenylalanine; G; guanosine; Gln; glutamine; Gly; glycine; Kb; kilobase(s); KDa; kilodalton(s); Km; kanamycin; LB; Luria–Bertani broth; Leu; leucine; Mb; megabase; O/N; overnight; PAGE; polyacrylamide gel electrophoresis; PCR; polymerase chain reaction; PIA; Pseudomonas; isolation agar; Pro; proline; Ser; serine; SDS; sodium dodecyl sulfate; T; thymidine; Tc; tetracycline; Tn; transposon; Trp or W; tryptophan; Tyr; tyrosine; V; valineExopolysaccharide; Alginate; Cystic fibrosis; P; algT; /; algU; Biosynthetic genes
Complement C3 gene polymorphism in renal transplantation (an Iranian experience)
by Najmeh Bazyar; Negar Azarpira; Saeid Reza Khatami; Hamid Galehdari; Heshmatolah Salahi (pp. 254-258).
The C3 component of complement has different roles in kidney disease and its local production in donor kidney may affect allograft function and rejection after organ transplantation. A single base substitution in c3 gene (rs2230199), defines two common allelic variants with different mobility on gel electrophoresis: S (Slow) and F (Fast). In order to evaluate the effect of this polymorphism on acute renal allograft rejection, one hundred samples of donor and recipients were collected and genotyping was done by PCR-RFLP method. The allelic frequencies were: C3S=0.791, C3F=0.209. There was no significant association between recipient's genotype and acute rejection ( p value<0.05). No correlation between donor genotype and acute rejection was also present. Patients were divided into four groups, according to the recipient and donor genotypes: SS recipients and FS or FF donor, SS recipient and donor, FF or FS recipient and SS donor and FS or FF recipient and donor. There was no significant difference in rate of acute rejection between groups. Although the results didn't find any association between C3 complement polymorphisms and acute allograft rejection, there was no acute rejection in FS or FF donors and SS recipient group.► No association between C3 polymorphisms and acute rejection was observed. ► There was no acute rejection in FS or FF donors and SS recipient group. ► The allelic frequencies were: C3S=0.791, C3F=0.209. ► No significant association between donor genotype and rejection episodes was detected.
Keywords: Complement system; Kidney allograft; Acute rejection
Isolation and characteristics of the melanocortin 1 receptor gene ( MC1R) in the Chinese yakow ( Bos grunniens× Bos taurus)
by Dongmei Xi; Min Wu; Yueyuan Fan; Yinqiang Huo; Jing Leng; Xiao Gou; Huaming Mao; Weidong Deng (pp. 259-263).
The Chinese yakow is the offspring of yak ( Bos grunniens) and Yellow cattle ( Bos taurus). The melanocortin 1receptor gene ( MC1R) plays a crucial role in determining coat colour of mammals. To investigate the relationship of polymorphism of the MC1R with coat colour in the Chinese yakow, the coding sequence (CDS) and the flanking region of MC1R were sequenced from 84 Chinese yakow samples and compared with the sequences of the MC1R from other bovid species. A fragment of 1134 base pair (bp) sequences including the full CDS (954bp) and parts of the 5′- and 3′-untranslated regions (162 and 18bp, respectively) of the Chineseyakow MC1R were obtained. A total of 13 single nucleotide polymorphisms (SNPs) including 4 SNPs (T-129C, A-127C, C-106T, G-1A) in the 5′-untranslated region and 9 SNPs (C201T, T206C, C340A, C375T, T663C, G714C, C870T, G871A and T890C) in the CDS were identified, revealing high genetic variability. Four novel SNPs including T206C, G714C, C870T and T890C, which have not been reported previously in bovid species, were retrieved. Within 9 coding SNPs, C201T, C375T, T663C and C870T were silent mutations, while T206C, C340A, G714C, G871A and T890C were mis-sense mutations, corresponding to amino acid changes p.L69P, p.Q114K, p.K238N, p.A291N and p.I297T, respectively. Amino acid sequences alignment showed a more than 96% similarity with other ruminates. However, three classical bovine MC1R loci the E D, E + and e were not retrieved in the Chinese yakow, indicating other genes or factors could be involved in affecting coat colour in this species.► The Chinese yakow MC1R was cloned and characterized for the first time. ► Four SNPs in the 5′-untranslated region and 9 SNPs in the CDS were identified. ► The three bovine MC1R E D, E + and e alleles were not detected in the Chinese yakow.
Keywords: Abbreviations; MC1R; melanocortin 1 receptor gene; CDS; the coding sequence; SNPs; single nucleotide polymorphisms; 5′-UTR; 5′-untranslated region; PCR; Polymerase chain reaction; Ho; gene homozygosity; He; gene heterozygosity; Ne; effective allele numbers; PIC; polymorphism information contentChinese yakow (; Bos grunniens; ×; Bos taurus; ); Melanocortin 1 receptor gene (; MC1R; ); Coat colour; Polymorphisms
RASSF1A Ala133Ser polymorphism is associated with increased susceptibility to hepatocellular carcinoma in a Turkish population
by Süleyman Bayram (pp. 264-269).
The tumor suppressor gene Ras association domain family 1 isoform A ( RASSF1A) regulates cell cycle regulation, apoptosis and microtubule stability and is inactivated by promoter hypermethylation at a high frequency in hepatocellular carcinoma (HCC). A guanine (G)/thymine (T) common single nucleotide polymorphism (SNP) at first position of codon 133 in RASSF1A gene determines an alanine (Ala) to serine (Ser) (Ala133Ser) amino acidic substitution which may alter cancer risk by influencing the function of RASSF1A protein.To determine the association of the RASSF1A Ala133Ser polymorphism with the risk of HCC development in a Turkish population, a hospital-based case–control study was designed consisting of 236 subjects with HCC and 236 cancer-free control subjects matched for age, gender, smoking and alcohol status. The genotype frequency of the RASSF1A Ala133Ser polymorphism was determined by using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay.Allele and genotype associations of RASSF1A Ala133Ser polymorphism with HCC susceptibility were observed in comparisons between the patient and control samples ( P<0.001). Risk of HCC development in this Turkish population was significantly increased in carriers of the Ser133 variant allele of Ala133Ser polymorphism (Ala/Ser and Ser/Ser genotypes) when compared with homozygote Ala/Ala genotype (OR=5.47, 95% CI=3.63–8.25, P=0.001).Because our results suggest for the first time that the Ser133 allele of RASSF1A Ala133Ser polymorphism may be a genetic susceptibility factor for HCC in the Turkish population, further independent studies are required to validate our findings in a larger series, as well as in patients of different ethnic origins.►RASSF1A protein plays important roles in tumorigenesis and metastasis of HCC ►I investigated the association of RASSF1A Ala133Ser polymorphism and risk of HCC. ►RASSF1A Ala133Ser polymorphism may be a genetic susceptibility factor for HCC.
Keywords: Abbreviations; RASSF1A; Ras association domain family 1; isoform A; SNP; Single nucleotide polymorphism; HCC; Hepatocellular carcinoma; CI; Confidence interval; OR; Odds ratioHepatocellular carcinoma; RASSF1A; Ala133Ser polymorphism; Genetic susceptibility; Case–control study
Renin–angiotensin system genes polymorphism in Egyptians with premature coronary artery disease
by Tarek A. Abd El-Aziz; Yousri M. Hussein; Randa H. Mohamed; Sally M. Shalaby (pp. 270-275).
Genetics polymorphism of the renin–angiotensin system (RAS) affects the pathogenesis of atherosclerosis and associated with coronary artery disease (CAD). We aimed to investigate the association between the RAS genes and premature CAD (PCAD) in Egyptians. 116 patients with PCAD, 114 patients with late onset CAD and 119 controls were included in the study. Angiotensin converting enzyme (ACE), angiotensin II receptor type 1 (ATR1) and angiotensinogen (AGT) genes polymorphisms were analyzed by polymerase chain reaction (PCR). We found that ACE DD, AGT TT and ATR1 CC increased the risk of PCAD by 2.7, 2.8 and 2.86 respectively). Smoking, hypertension, diabetes, total cholesterol, triglycerides and LDL cholesterol were independent risk factors for the development of PCAD. We conclude that the ACE DD, AGT TT and ATR1 CC genotypes may increase the susceptibility of an individual to have PCAD. The coexistence of CAD risk factors with these risky RAS genotypes may lead to the development of PCAD in Egyptian patients.► ACE DD genotypes may increase the susceptibility of an individual to have PCAD. ► AGT TT genotypes may increase the susceptibility of an individual to have PCAD. ► ATR1 CC genotypes may increase the susceptibility of an individual to have PCAD. ► Coexistence of CAD risk factors with RAS genotypes lead to the development of PCAD.
Keywords: RAS; Genetic polymorphism; Premature CAD
Rapid genotyping of APOA5 −1131T>C polymorphism using high resolution melting analysis with unlabeled probes
by Song-mei Liu; Feng-xia Xu; Fan Shen; Yan Xie (pp. 276-279).
The APOA5 −1131 T/C polymorphism (rs662799) exhibits a very strong association with elevated TG levels in different racial groups. High resolution melting (HRM) analysis with the use of unlabeled probes has shown to be a convenient and reliable tool to genotyping, but not yet been used for detecting rs662799 polymorphism. We applied the unlabeled probe HRM analysis and direct DNA sequencing to assay the −1131T>C SNP in 130 cases DNA samples blindly. This HRM analysis can be completed in <3min for each sample. The two melting peaks were displayed at 66.1±0.4°C for CC homozygote and 68.7±0.2°C for TT homozygote; TC heterozygote showed the both melting peaks. The genotyping results by HRM method were completely concordant with direct DNA sequencing. The distribution of CC, TC, and TT genotypes for the −1131T>C SNP was 9.2, 49.2, and 41.5%, respectively. This assay was sensitive enough to detect C allele down to 20% and 10% for T allele. The limit of detection for C and T allele was 6.2 and 2.5ng/μL DNA, respectively. The developed unlabeled probe HRM method provides an alternative mean to detect ApoA5 −1131T>C SNP rapidly and accurately.► We developed an unlabeled probe HRM analysis to assay the APOA5 −1131 T/C polymorphism (rs662799). ► This assay was sensitive enough to detect C allele down to 20% and 10% for T allele. ► The limit of detection for C and T allele was 6.2 and 2.5ng/μL DNA, respectively.
Keywords: Abbreviations; ApoA5; apolipoprotein A5; PCR; polymerase chain reaction; HRM; high resolution melting; SNP; single nucleotide polymorphism; Tm; melting temperatureHigh resolution melting; Apolipoprotein A5; Single nucleotide polymorphism; Genotyping
DNA adenine methyltransferase (Dam) controls the expression of the cytotoxic enterotoxin ( act) gene of Aeromonas hydrophila via tRNA modifying enzyme-glucose-inhibited division protein (GidA)
by Tatiana E. Erova; Valeri G. Kosykh; Jian Sha; Ashok K. Chopra (pp. 280-287).
Aeromonas hydrophila is both a human and animal pathogen, and the cytotoxic enterotoxin (Act) is a crucial virulence factor of this bacterium because of its associated hemolytic, cytotoxic, and enterotoxic activities. Previously, to define the role of some regulatory genes in modulating Act production, we showed that deletion of a glucose-inhibited division gene ( gidA) encoding tRNA methylase reduced Act levels, while overproduction of DNA adenine methyltransferase (Dam) led to a concomitant increase in Act-associated biological activities of a diarrheal isolate SSU of A. hydrophila. Importantly, there are multiple GATC binding sites for Dam within an upstream sequence of the gidA gene and one such target site in the act gene upstream region. We showed the dam gene to be essential for the viability of A. hydrophila SSU, and, therefore, to better understand the interaction of the encoding genes, Dam and GidA, in act gene regulation, we constructed a gidA in-frame deletion mutant of Escherichia coli GM28 ( dam +) and GM33 (∆ dam) strains. We then tested the expressional activity of the act and gidA genes by using a promoterless pGlow-TOPO vector containing a reporter green fluorescent protein (GFP). Our data indicated that in GidA+ strains of E. coli, constitutive methylation of the GATC site(s) by Dam negatively regulated act and gidA gene expression as measured by GFP production. However, in the ∆ gidA strains, irrespective of the presence or absence of constitutively active Dam, we did not observe any alteration in the expression of the act gene signifying the role of GidA in positively regulating Act production. To determine the exact mechanism of how Dam and GidA influence Act, a real-time quantitative PCR (RT-qPCR) assay was performed. The analysis indicated an increase in gidA and act gene expression in the A. hydrophila Dam-overproducing strain, and these data matched with Act production in the E. coli GM28 strain. Thus, the extent of DNA methylation caused by constitutive versus overproduction of Dam, as well as possible conformation of DNA influence the expression of act and gidA genes in A. hydrophila SSU. Our results indicate that the act gene is under the control of both Dam and GidA modification methylases, and Dam regulates Act production via GidA.► Cytotoxic enterotoxin (Act) of A. hydrophila (Ah) is regulated by GidA and Dam. ► Both Dam and GidA are needed for Act production in E. coli (Ec). ► gidA mutants produce similar level of Act (as GFP) in Ec irrespective of dam status. ► Methylation of gidA upstream region by constitutive Dam decreases Act production. ► Hypermethylation of GidA by Dam overproduction increases Act production in Ah. ► Cross-talk between Act, GidA, and Dam governs virulence gene expression.
Keywords: Abbreviations; Act; Aeromonas; cytotoxic enterotoxin; Dam; DNA adenine methyltransferase; tRNA; transfer RNA; GidA; glucose-inhibited division A protein; GATC site; guanine-adenine-thymine-cytosine site; RT-qPCR; real-time quantitative polymerase chain reaction; GFP; green fluorescent protein; GAPDH; glyceraldehyde-3-phosphate dehydrogenase; DPBS; Dulbecco's phosphate-buffered saline; FAD; Flavin adenine dinucleotide; OriC; origin of replication; Ap; Ampicillin; Cm; Chloramphenicol; Rif; Rifampin; gDNA; genomic DNA; RFU; relative fluorescence units; FRT; flippase [FLP] recognition target; Cm; s; chloramphenicol sensitive; Ap; s; ampicillin sensitive; pap; pyelonephritis-associated pili; seqA; gene; sequestration A geneGATC Dam target sites; Promoter activity; tRNA uridine 5 carboxymethylaminomethyl modification enzyme
The complete mitochondrial genome of Microtus fortis calamorum (Arvicolinae, Rodentia) and its phylogenetic analysis
by Xianhuan Jiang; Jun Gao; Liju Ni; Jianhua Hu; Kai Li; Fengping Sun; Jianyun Xie; Xiong Bo; Chen Gao; Junhua Xiao; Yuxun Zhou (pp. 288-295).
Microtus fortis is a special resource of rodent in China. It is a promising experimental animal model for the study on the mechanism of Schistosome japonicum resistance. The first complete mitochondrial genome sequence for Microtus fortis calamorum, a subspecies of M. fortis (Arvicolinae, Rodentia), was reported in this study. The mitochondrial genome sequence of M. f. calamorum (Genbank: JF261175) showed a typical vertebrate pattern with 13 protein coding genes, 2 ribosomal RNAs, 22 transfer RNAs and one major noncoding region (CR region).The extended termination associated sequences (ETAS-1 and ETAS-2) and conserved sequence block 1 (CSB-1) were found in the CR region. The putative origin of replication for the light strand (OL) of M. f. calamorum was 35bp long and showed high conservation in stem and adjacent sequences, but the difference existed in the loop region among three species of genus Microtus. In order to investigate the phylogenetic position of M. f. calamorum, the phylogenetic trees (Maximum likelihood and Bayesian methods) were constructed based on 12 protein-coding genes (except for ND6 gene) on H strand from 16 rodent species. M. f. calamorum was classified into genus Microtus, Arvcicolinae for the highly phylogenetic relationship with Microtus kikuchii (Taiwan vole). Further phylogenetic analysis results based on the cytochrome b gene ranged from M. f. calamorum to one of the subspecies of M. fortis, which formed a sister group of Microtus middendorfii in the genus Microtus.The organization of the M. f. calamorum mitochondrial genome. The tRNAs were denoted using single letter amino acid code. The protein coding genes were indicated as follows: ND1-6, NADH dehydrogenase subunits 1–6; COX1–3, cytochrome c oxidase subunits 1–3; ATP6 and ATP8, ATPase subunits 6 and 8; Cyt b, Cytochrome b; and CR, control region.Display Omitted► The first mitochondrial genome of M. f. calamorum was reported in this study. ► The mitochondrial genome of M. f. calamorum showed a typical vertebrate pattern. ► M. f. calamorum show highly phylogenetic relationship with Microtus kikuchii. ► M. f. calamorum formed a sister group with Microtus middendorfii in the genus Microtus.
Keywords: Abbreviations; M. f. calamorum; Microtus fortis calamorum; M. fortis; Microtus fortis; mtDNA; mitochondrial DNA; atp6 and atp8 genes; ATP synthase subunits 6 and 8; Cyt b gene; cytochrome b; cox1–3 genes; subunits 1–3 of cytochrome c oxidase; ND1–6 and ND4L; NADH dehydrogenase subunits 1–6 and 4L; 12S rRNA and 16S rRNA genes; large and small subunits of ribosomal RNA; tRNA; transfer RNA; DHU; dihydrouridine loop; CR; Control region; PCR; polymerase chain reaction; TΨC; pseudouridine loop; kb; kilobase; bp; base pair; ML; Maximum likelihood; NJ; Neighbor-joining; GTR; +; G; +; I; General time reversible model with a gamma distributed shape parameter and the proportion of invariable sites; Mya; million years ago Microtus fortis calamorum; Mitochondrial genome; Phylogenetic relationship
A case of del(13)(q14.2)(q31.3) associated with hypothyroidism, hypertriglyceridemia, hypercholesterolemia and total ophthalmoplegia
by Baris Malbora; Cihan Meral; Nihan Malbora; Deniz Sunnetci; Naci Cine; Hakan Savli (pp. 296-299).
13q deletion syndrome is caused by the absence of a portion of the long arm of chromosome 13. This syndrome is a rare condition characterized by a wide range of clinical findings. Phenotype varies with the location and size of the deletion. We report a female dizygotic twin with a proximal deletion of 13q and failure to thrive, hypotonia, and multiple anomalies included pytosis and total ophthalmology at right side, strabismus at left, bilateral iris heterochromia and telecantus. She had a broad nasal bridge with flat philtrum, micrognathia and antevert ear lobes. Her umbilicus had vanished. Her left coxa was dislocated and left toes were overlapped. She was also found to have hypertriglyceridemia, hypercholesterolemia, and hypothyroidism. Chromosome analysis showed a proximal deletion of chromosome 13 [karyotype 46,XX,del(13) (q14.2q31.3)] which was confirmed by high-resolution microarray based comparative genomic hybridization.The described patient is unique among similar rare cases with different deletion breakpoints. It is the first case of 13q14.2q31.3 deletion where the breakpoints are clearly defined, indicating the importance of detailed clinical description and high-resolution genomic analysis for characterization of rare genetic syndromes.► New findings in 13q deletion syndrome are detected. ► We detected a new eye finding, total ophthalmoplegy in this syndrome. ► It is important to keep in mind that hypothyroidism and lipid abnormalities may be present.
Keywords: Abbreviations; BAC; bacterial artificial chromosome; CGH; comparative genomic hybridization; del; deletion; DNA; deoxyribo nucleic acid; EEG; electroencephalography; VLDL; very low density lioproteinChromosome 13q deletion; Ophthalmoplegia; Hypertriglyceridemia; Hypercholesterolemia; Hypothyroidism; Array-CHG
Involvement of p53 in gemcitabine mediated cytotoxicity and radiosensitivity in breast cancer cell lines
by Sameer D. Salem; Faisal M. Abou-Tarboush; Nadeem M. Saeed; Waheeb D. Al-Qadasi; M. Abul Farah; Muneera Al-Buhairi; Najla Al-Harbi; Ibrahim Alhazza; Ghazi Alsbeih (pp. 300-307).
Gemcitabine (2′,2′-difluoro-2′-deoxycytidine; dFdCyd) is one of the anti-metabolites drugs that target DNA replication. We evaluated dFdCyd cytotoxicity and its radiosensitizing ability in human breast cancer cell lines, MCF-7 (wild-type p53) and MDA-MB-231 (mutant-type p53) along with normal mammary epithelial cell line (MCF-12) for comparison. Radiosensitivity and cytotoxicity were measured by the clonogenic survival assays. DNA DSBs was studied by Pulse Field Gel Electrophoresis (PFGE) and cell cycle distribution was analyzed by flow cytometry. MDA-MB-231 cells were the most sensitive to the cytotoxicity of dFdCyd (IC50 5nM) then MCF-7 (IC50 10nM), whereas MCF-12 cells were the most resistant to the cytotoxicity of dFdCyd (IC50 70nM). MCF-12 and MCF-7 cell lines did not show any radiosensitization to dFdCyd, whereas the MDA-MB-231 cells showed significantly increased radioresistant to dFdCyd at equimolar concentration (p=0.002) and at IC50 concentration (p<0.001). The DNA double strand breaks (DSBs) repair showed that dFdCyd neither increases DNA DSBs nor decreases the rate of their repair in MCF-12 and MCF-7 cell lines, while the same treatment in MDA-MB-231 cell line led to decrease the rate of DSBs or increase the rate of DNA repair (p=0.034). Therefore, dFdCyd is a cytotoxic agent, especially in the cancer cells irrespective of having wild-type or mutated p53 protein, but it is not effective as radiosensitizer in the cell lines used in this study. dFdCyd combined with radiation reduces the efficacy of chemo-radiotherapy in p53 mutated cells. Therefore, p53-mutated cancer could be a counter-indication for radiation–gemcitabine combined treatment.► Gemcitabine is a potent cytotoxic agent for cancer cell lines. ► Gemcitabine reduces the efficacy of chemo-radiotherapy in p53 mutated tumors. ► Gemcitabine neither increases DNA DSBs nor decreases its repair in cancer cells.
Keywords: Abbreviations; dFdCyd; gemcitabine (2′,2′-difluoro-2′-deoxycytidine); DSB; double strand break; PFGE; pulsed field gel electrophoresis; PBS; phosphate buffered saline; Na; 2; EDTA; sodium ethylenediamine tetraacetic acid; TBE; Tris-Borate-EDTA buffer; MID; mean inactivation dose; SF2; surviving fraction at 2; Gy; PI; propidium iodide; IC; inhibitory concentrationGemcitabine; p53; DNA double-strand breaks; Cytotoxicity; Radiosensitivity; Breast cancer
Congenital heart defect and mental retardation in a patient with a 13q33.1-34 deletion
by Can Huang; Yi-Feng Yang; Ni Yin; Jin-Lan Chen; Jian Wang; Hong Zhang; Zhi-Ping Tan (pp. 308-310).
13q deletion syndrome is a rare genetic disorder caused by deletions of the long arm of chromosome 13. Patients with 13q deletion display a variety of phenotypic features. We describe a one-year-old female patient with congenital heart defects (CHD), facial anomalies, development and mental retardation. We identified a 12.75Mb deletion in chromosome region 13q33.1-34 with high resolution SNP Array (Human660W-Quad, Illumina, USA). This chromosome region contains about 55 genes, including EFNB2, ERCC5, VGCNL1, F7, and F10. Comparing our findings with previously reported 13q deletion patients with congenital heart defects, we propose that the 13q33.1-34 deletion region might contain key gene(s) associated with cardiac development. Our study also identified a subclinical deficiency of Factors VII and X in our patient with Group 3 of 13q deletion syndrome.► High resolution SNP array identified a 13q deletion in a patient with CHD and MR. ► Deficiency of Factors VII and X in the proband with 13q deletion syndrome. ► 13q33.1-34 region might be one of the regions responsible for CHD. ► A locus for CHD was suggested to a minimal deletion of 6Mb in13q33.1-34 region.
Keywords: Abbreviations; CNV; copy number variation; SNP; single-nucleotide polymorphism; CHD; congenital heart defects; Hg18; human genome; VSD; ventricular septal defect; ASD; arterial septal defect; DGV; Database of Genomic Variants; TOF; tetralogy of Fallot13q deletion syndrome; Congenital heart defect; Copy number variation; Microdeletion; Single-nucleotide polymorphism array
Association study of AGER gene polymorphism and hypertension in Han Chinese population
by Song Yang; Hairu Wang; Yichun Yang; Wen Wang; Jiandong Jiang; Xianghai Zhao; Qinglian Du; Xuecai Wang; Yingshui Yao; Hongbing Shen; Chong Shen; Yanping Zhao (pp. 311-316).
Advanced glycation end products (AGEs) are produced by non-enzymatic glycation or glycoxidation of proteins, lipids and nucleic acids. The bond of AGEs and the receptor of AGE (AGER) in a pro-oxidant environment could induce immune and inflammation reaction involved in progress of microvascular disease. Accumulated evidence warrant further study on AGE–AGER pathway and genetic susceptibility to hypertension (HT).We designed a two-stage association study to evaluate the association of AGER polymorphism and HT. In stage 1, seven tagSNPs were tested in 524 cases and 531 controls and the significant SNPs ( P<0.05) would enter into stage 2 including 807 cases and 869 controls. Furthermore, joint analysis was performed for all 2731 subjects including 1331 cases and 1400 controls, and meta-analysis was applied to evaluate combined estimations from the subgroups of stage 1 and stage 2.In stage 1, rs204994 had significant association with HT ( P<0.05) and enter stage 2. Neither joint analysis nor meta-analysis found statistical association of rs204994 with HT after adjusted for the covariates in the whole population. However, further stratification analysis found that rs204994 was significantly associated with HT in <50years and ≥50years groups, ORs (95%CI) of dominant model were 1.623 (1.054–2.500) and 0.721 (0.546–0.952) respectively. No significant correlation was found between blood pressure and the polymorphisms of rs204994.Our data suggests that age might modulate the genetic effects of variation of rs204994 in AGER on HT and further replications in other populations and functional studies should be warranted.► A allele of rs204994 presented significant risk effect on HT in <50years group. ► A allele of rs204994 showed protective effect on HT in ≥50years group reversely. ► No significant effect of rs204994 was found on blood pressure in this population.
Keywords: Abbreviations; AGEs; advanced glycation end products; AGER; receptor for AGE; Ang; angiotensin; BMI; body mass index; χ; 2; Chi-square; CHB; Chinese population in Beijing; CAD; coronary artery disease; DBP; diastolic blood pressure; esRAGE; endogenous secretory RAGE; EDTA; ethylenediamine tetraacetic acid; GLM; general linear model; GLU; glucose; HWE; Hardy–Weinberg equilibrium; HDL-C; high density lipoprotein cholesterol; HMGB1; high-mobility group box-1; HT; hypertension; LD; linkage disequilibrium; MAF; minor allele frequency; NF-κB; nuclear factor kappa-B; OR; odds ratio; CI; 95% confidence; PBX2; interval pre-B-cell leukemia homeobox 2; PCR-RFLP; polymerase chain reaction and restriction fragment length polymorphism; P; probability; RAS; renin–angiotensin system; ROS; reactive oxygen species; SNPs; single nucleotide polymorphisms; SHRSP; spontaneously hypertensive rats; SPSS; Statistical Product and Service Solutions; SBP; systolic blood pressure; tagSNPs; tagger SNPs; TLR; toll-like receptor; TFBS; transcript factor banding site; TGR; transgenic rat; TG; triglycerides; TC; total cholesterolAdvanced glycation end products; Receptor of advanced glycation end products; Hypertension; Association study
Polymorphisms of glutathione S-transferases M1, T1, P1 and A1 genes in the Tunisian population: An intra and interethnic comparative approach
by Ghada Ben Salah; Fakhri Kallabi; Sirine Maatoug; Emna Mkaouar-Rebai; Amine Fourati; Faiza Fakhfakh; Hamadi Ayadi; Hassen Kamoun (pp. 317-322).
Genetic polymorphisms in glutathione S-transferases (GSTs) genes might influence the detoxification activities of the enzymes predisposing individuals to cancer risk. Owing to the presence of these genetic variants, inter-individual and ethnic differences in GSTs detoxification capacity have been observed in various populations. Therefore, the present study was performed to determine the prevalence GSTM1*0/*0, GSTT1*0/*0, GSTP1 Ile105Val, and GSTA1*A/*B polymorphisms in 154 healthy individuals from South Tunisia, and to compare them with those observed in North and Centre Tunisian populations and other ethnic groups. GSTM1 and GSTT1 polymorphisms were analyzed by a Multiplex-PCR approach, whereas GSTP1 and GSTA1 polymorphisms were examined by PCR-RFLP. The frequencies of GSTM1*0/*0 and GSTT1*0/*0 genotypes were 53.9% and 27.9%, respectively. The genotype distribution of GSTP1 was 47.4% (Ile/Ile), 40.9% (Ile/Val), and 11.7% (Val/Val). For GSTA1, the genotype distribution was 24.7% (*A/*A), 53.9% (*A/*B), and 21.4% (*B/*B). The combined genotypes distribution of GSTM1, GSTT1, GSTP1 and GSTA1 polymorphisms showed that thirty one of the 36 possible genotypes were present in our population; eight of them have a frequency greater than 5%. To the best of our knowledge, this is the first report of GSTs polymorphisms in South Tunisian population. Our findings demonstrate the impact of ethnicity and reveal a characteristic pattern for Tunisian population. The molecular studies in these enzymes provide basis for further epidemiological investigations in the population where these functional polymorphisms alter therapeutic response and act as susceptibility markers for various clinical conditions.► The genetic polymorphisms in GSTs genes might influence their activities. ► Intra/interethnic differences have been known in the prevalence of GSTs polymorphisms. ► Southern Tunisians were genotyped for the major four GSTs polymorphisms. ► Results show a characteristic pattern for Southern Tunisia population. ► The data may provide a basic database for the future clinical and genetic studies.
Keywords: Abbreviations; GSTs; Glutathione S-transferases; GSTM1; Glutathione S-transferase M1; GSTT1; Glutathione S-transferase T1; GSTP1; Glutathione S-transferase P1; GSTA1; Glutathione S-transferase A1; PCR; Polymerase Chain Reaction; RFLP; Restriction fragment Length Polymorphism; DNA; Deoxyribonucleic acid; SNP; Single-nucleotide polymorphismGlutathione S-transferase; Intra/interethnic variability; Combined genotypes; Tunisian population
Expression and characterization of a recombinant Cry1Ac crystal protein fused with an insect-specific neurotoxin ω-ACTX-Hv1a in Bacillus thuringiensis
by W.P. Li; L.Q. Xia; X.Z. Ding; Y. Lv; Y.S. Luo; S.B. Hu; J. Yin; F. Yan (pp. 323-327).
In order to assess possible enhancement of biopesticide activity, the fusion gene of crystal protein gene cry1Ac with the insect-specific neurotoxin ω-ACTX-Hv1a gene and egfp was expressed in Bacillus thuringiensis acrystalliferous strain Cry-B under the control of the native gene expression system. The fusion recombinant Cry-B(1Ac-ACTX-EGFP) generally produced two or three small crystal-like inclusion bodies in each cell and the GFP signal could be clearly observed. A 166kDa full-length fusion protein was identified by immunoblot analysis. Virulence of the fusion inclusions was at least fivefold higher toward larvae of Spodoptera exigua. These results demonstrated that a foreign protein could be expressed and accumulate as parasporal inclusions in B. thuringiensis by C-terminal fusion with the native endotoxin while retaining partial insecticidal activity.► The fusion protein Cry1Ac-ω-ACTX-Hv1a-EGFP was successfully expressed in B. t. ► Heterogeneous toxins formed parasporal inclusion bodies together with endotoxins. ► Virulence of the fusion inclusions was significantly improved against S. exigua. ► It is a promising approach for potent biopesticides with lower resistance potential.
Keywords: Abbreviations; AFM; atomic force microscope; bp; base pair; B. t.; Bacillus thuringiensis; E. coli; Escherichia coli; EGFP; enhanced green fluorescent protein; H. armigera; Helicoverpa armigera; kDa; kiloDalton; LC; 50; 50% lethal concentration; PAGE; polyacrylamide gel electrophoresis; PCR; polymerase chain reaction; PVDF; poly(vinylidene fluoride); rpm; revolutions per minute; S. exigua; Spodoptera exigua; SDS; sodium dodecyl sulfate; STAB-SD; Stabilized-Shine-Dalgarno (sequence); Subsp.; subspecies; VGCCs; voltage-gated calcium channelsFusion gene; Crystallization; Neurotoxin; Insecticidal activity
16q22.1 microdeletion detected by array-CGH in a family with mental retardation and lobular breast cancer
by Chiara Palka Bayard de Volo; Melissa Alfonsi; Valentina Gatta; Antonio Novelli; Laura Bernardini; Donatella Fantasia; Ivana Antonucci; Domenico Angelucci; Robert Zori; Liborio Stuppia; Francesco Chiarelli; Giuseppe Calabrese (pp. 328-331).
We describe the case of a boy with psychomotor delay and dysmorphic features, with a germline 16q22.1 microdeletion identified by array-CGH. The deletion spans 0.24Mb and encompasses three genes (ZFP90, CDH3 and CDH1). The deletion has been demonstrated to be inherited from his mother who was affected by lobular breast cancer (LBC) without any other apparently phenotypic features. We suppose that the microdeletion, in particular ZFP90 which is cerebrally expressed, is causative for the boy's phenotype. Mental retardation in the affected boy can recognize several mechanisms such as variable expressivity, non-penetrance, multifactorial/polygenic inheritance, recessive inheritance, a second rearrangement event and epigenetics. Furthermore, we suggest that the deletion of the CDH1, a tumor suppressor gene, involved in hereditary diffuse gastric cancer (HDGC) and LBC predisposed the mother to the carcinoma.► First case of 16q22.1 microdeletion associated with mental retardation. ► The deletion encompass three genes: ZFP90, CDH1 and CDH3. ► ZFP90 is a candidate gene for mental retardation. ► CDH1 deletion predisposed the mother to the breast cancer.
Keywords: Abbreviations; ZFP90; Zinc finger protein 90; CDH3; Cadherin-3; CDH1; Cadherin-1; LBC; lobular breast cancer; HDGC; hereditary diffuse gastric cancer; array-CGH; Array-comparative genomic hybridization; EEM; Ectodermal dysplasia, Ectrodactyly and Macular dystrophy; GC; gastric cancer; IGCLC; International Gastric Cancer Linkage Consortium; MRI; magnetic resonance imaging; BRCA1; Breast cancer 1; BRCA2; Breast cancer 2; PCR; polymerase chain reaction; FISH; Fluorescence in situ hybridization; LCIS; lobular carcinoma; in situ; LSAB; Labeled Streptavidin Biotin; UPD; Uniparental disomy; CNV; Copy number variant; ACD; adrenocortical dysplasiaZFP90; CDH1; Array-CGH
Genotype/phenotype of 6 Chinese cases with Niemann–Pick disease type C
by Hui Xiong; Katsumi Higaki; Cui-jie Wei; Xin-Hua Bao; Yue-Hua Zhang; Na Fu; Jiong Qin; Kaori Adachi; Yumiko Kumura; Haruaki Ninomiya; Eiji Nanba; Xi-Ru Wu (pp. 332-335).
Niemann–Pick disease type C (NP-C), caused by mutations of either NPC1 or NPC2 gene, is an inherited lysosomal lipid storage disorder that is difficult to be diagnosed and treated. NP-C is rarely reported in China and so far very few literatures are available for Chinese clinical workers. To better characterize this disease in China and improve genetic counseling, mutational analyses of NPC1 gene were carried out in 6 unrelated Chinese patients.Clinical data of the probands from 2007 to 2010 were collected and analyzed. All exons of NPC1 were analyzed by direct sequencing.The six cases, four males and two females, included three cases of late infantile subtype and three cases of juvenile subtype. Case one and case six had siblings who suffered from the same disease. The onset of clinical symptoms varied from three to ten years old, and they included progressive cognitive and language impairment, and motion retrogradation. All were caught by focal or generalized seizures from one to four years after the onset. Vertical supranuclear gaze palsy, dysarthria, dysphagia, internal rotation and adduction of bilateral hands and splenomegaly occurred gradually during the disease progression. Five patients had laughter-cataplexy. MRI indicated mild brain atrophy. Sea blue cells and Niemann–Pick cells were presented in bone marrow smears. Activity of acid sphingomyelinase was normal or only slightly lower than controls. Supporting and symptomatic treatments could improve some of the clinical signs. We identified 10 different NPC1 mutations were identified in 12/12 alleles, 3 of which are described for the first time. All mutations were missense mutations, which located throughout the gene with five clustering in the cysteine-rich luminal domain. Homozygous mutation of S865L correlated with a relatively severe juvenile neurological form.NP-C is a rare autosomal recessive lysosomal storage disease that affects intellectual development of children, causing dementia, vegetative state and eventual death. The awareness of NP-C should be raised in the Chinese population. The typical clinical features of this disease include vertical supranuclear gaze palsy, seizures and cataplexy. Laboratory features include the presence of sea blue cells and Niemann–Pick cells in bone marrow smears. NPC1 mutation can be identified in most of these patients and most of them are missense mutations.► We performed clinical and genetic analyses in six Chinese NP-C patients. ► We identified 10 different NPC1 mutations with three novel mutations. ► Our study will be helpful for diagnosis of Chinese NP-C patients.
Keywords: Abbreviations; NP-C; Niemann–Pick disease type C; NPC1; Niemann–Pick disease type C1; NPC2; Niemann–Pick disease type C2; PCR; polymerase chain reaction; C-WISC; Wechsler Intelligence Scale for Children; MRI; magnetic resonance imaging; EEG; electroencephalogram; LDL; low-density lipoprotein; HDL; high-density lipoproteinNiemann–Pick disease type C; Sea blue cells; Vertical supranuclear gaze palsy; NPC1
Wiley Announces the 10th edition of the Wiley Registry of Mass Spectral Data
November 22, 2013
2-day In-person Seminar on Mastering the HIPAA Privacy, Security, and Breach Notification Rules : Coping with Rule Changes, Managing Incidents, Preparing for Audits, and Avoiding Penalties
November 14, 2013
Dolomite to showcase novel microfluidic solutions at MicroTAS 2013
October 17, 2013
Dolomite's new Membrane Chip Interface advances multiple microfluidic applications
October 10, 2013
Looking for the next generation in microfluidics
October 1, 2013