isolation cloning and expression of genes

... Cycling Assay 81 Discussions 4.1 Isolation and Characterization of Conidial NADase from N crassa 83 4.2 Identification of NADase Sequence 85 4.3 Cloning and Expression of Gi|85106032 86 4.4 NAD+ Cycling ... blot analysis of supernatant of intracellular expression and secretion expression recombinant proteins expressed by P pastoris 75 3.19 Time course of intracellularly expressed and secreted recombinant ... and kanr Intracellular Expression cassette bounded by BamHI and BglII sites for generation of multicopy expression vector Multiple cloning sites for insertion of foreign genes; G418 selection...

Ngày tải lên: 08/11/2015, 17:15

... other hand, it was inhibited by high concentrations of azide, an inhibitor of CD39 and most ecto-ATPDases [29–31], and high concentrations of fluoride, vanadate, and pyrophosphate, inhibitors of the ... purification of the chicken liver ecto-ATPDase, its enzymatic properties and localization, cloning of the full-length cDNA and its expression EXPERIMENTAL PROCEDURES Materials The production and characterization ... absence and presence of the indicated concentrations of sodium azide.MgATP hydrolysis at pH 7.4 (s) and 6.4 (d); MgADP hydrolysis at pH 7.4 (h) and 6.4 (j) inhibition by azide was of the mixed and...

Ngày tải lên: 22/02/2014, 04:20

... well as a genomic DNA clone were isolated and sequenced, and the expression and modulation of this molecule was studied Results Cloning and characterization of rainbow trout IL-11 A cDNA clone containing ... infection, and a strong product was present in samples prepared at 24 and 48 h after infection (Fig 4) Expression and modulation of trout IL-11 in cell lines The constitutive expression of IL-11 ... myofibroblasts, IL-b and TGF-b are potent inducers of IL-11 expression at both the mRNA and protein levels [8] Trout RTS-11 cells show constitutive expression of TGF-b mRNA, and IL-1b but not TGF-b...

Ngày tải lên: 07/03/2014, 16:20

... of peak area ratios between m/z 426 and m/z 372 ions (C27D8,14/5acholestane) or m/z 428 and m/z 372 ions (C27D8/5acholestane) at the expected retention time RESULTS AND DISCUSSION Isolation of ... pCR2.1 vector, and sequenced on both strands The cloned cDNA was 1370 bp long and contained an ORF of 1257 bp, encoding a protein of 418 amino acids with a calculated molecular mass of 46 751 Da ... dithiothreitol, 40 UámL)1 of RNase inhibitor, 2.5 mM dNTP, 0.2 mM oligo-dT 15±18mer, and 200 U of M-MLV reverse transcriptase First-strand synthesis was performed at 42 °C for 45 and then the enzyme...

Ngày tải lên: 08/03/2014, 16:20

... speculated that the physical role of MeJA is to mobilize JA [19] The proof of this hypothesis might come from a more detailed understanding of how plants form MeJA from JA, and vice versa A recent study ... cycles of 30 s at 94 °C, at 40 °C and at 72 °C, followed by 30 cycles of 30 s at 94 °C, at 45 °C and at 72 °C Gel electrophoresis, DNA elution and modification was carried out according to standard ... Bacterial expression and purification of tomato MJE To obtain unequivocal evidence of the identity of the cloned sequence, MJE cDNA was subcloned into a bacterial vector for heterologous expression...

Ngày tải lên: 16/03/2014, 18:20

... reports on the cloning and expression of two AKR genes from D purpurea Both proteins reduce the ketone group of steroid structures but they are not active on the D4-double bond of the steroids ... of the steroids assayed It is worth noting that this is the first report for such steroid activity from a plant AKR enzyme Expression of DpAR genes The size of the DpAR transcripts and their expression ... enzymatic activity on steroids, the organ-specific and developmentally regulated expression of the genes, and the specific biosynthesis and accumulation of cardenolides in mature Digitalis leaves Once...

Ngày tải lên: 18/03/2014, 01:20

... relative expression value of one The expression of both splice variants of L-fucokinase was found to be relatively high in brain, ovary, testis and kidney In spleen, heart and lung the expression ... time quantitative PCR assay [28] to detect the expression levels of pyrophosphorylase and long and short splice variants of fucokinase Primers and probes were designed using the PRIMEREXPRESS ... L-fucokinase and pyrophosphorylase The primer and probe sequences and their positions in the mRNA sequence, for GDP-L-fucose pyrophosphorylase and the short and long splice variants of L-fucokinase,...

Ngày tải lên: 30/03/2014, 13:20

... 3.1 Homology of PfMCA1 39 3.2 Expression of PfMCA1 and YCA1 protein in yeast 40 3.3 Optimization of protein expression 42 3.4 Expression of optimized PfMCA1 and YCA1 amplified ... discarded and the cell pellet was washed with ml of TE buffer The cell pellet was resuspended in 0.5 ml of TE buffer, and 20 µl of 10% SDS, 10 µl of proteinase K (from a stock of 10 µg/µl solution), and ... 4.7 4.8 PfMCA1 expression in E coli 69 PfMCA1 expression in T brucei 71 Over -expression of YCA1 73 Amplification of PfMCA1 from RNA 74 Expression of PfMCA1 variants...

Ngày tải lên: 03/10/2015, 20:57

... Transcriptional regulation of the gonadotropin subunit genes The regulation of gonadotropin genes comprises basal gene expression which targets these genes specifically to the gonadotropes, and hormonal stimulation ... levels of FSH and LH LH FSH Mar Apr May Jun July Aug vitellogenesis Sep Oct Nov ovulation Figure 1: Plasma levels of FSH and LH in coho salmon during reproductive maturation (modified from Dickhoff ... Dickhoff and Swanson, 1990) The expression and pulsatile secretion of these hormones are temporally regulated throughout the reproductive cycle and are subject to the complex control of hypothalamic...

Ngày tải lên: 08/11/2015, 17:15

... protein products of highly overexpressing (NaCl-induced) genes after ATB-1 and HDZ3 showing EST redundancy of 0.89 and 1.17%, respectively In addition, there was overexpression of genes of two other ... although cloning and sequencing of the cDNAs from the forward subtracted SSH cDNA library did not show any presence of EST of the gene Activity assay of catalase and P5CS The activity of catalase and ... discovery of more such genes, particularly that of the low abundance proteins, on continued cloning and sequencing of the forward SSH cDNA library Moreover, more than 30% of the unigenes Page 16 of...

Ngày tải lên: 12/08/2014, 03:20

... growth and mortality of insect pests, Plutella xylostella (PX) and Spodoptera litura (SL).17th International Congress of Biochemistry and Molecular Biology Annual Meeting of the American Society of ... primers and the purified total MCo DNA as template The conditions for this experiment were established as: 150ng of each primer; 200àM of dNTPs ;400ng of the purified total MCo DNA Cloning of the ... Sequence of recombinant plasmid DNA containing TI gene fragment Expression of recombinant MCoTI-II (ReMCoTI-II) The expression conditions such as ampicillin and IPTG concentrations, temperature and...

Ngày tải lên: 12/02/2014, 10:20

... GCN2-independent regulation of gene expression Our results show that rapamycin, an inhibitor of mTORC1, regulates the expression of a set of genes almost as large as the set of genes regulated by amino ... treatment and amino acid starvation had similar effects on the expression of 12 genes and opposite effects on the regulation of eight genes These results suggest that rapamycin inhibition of TORC1 ... behavior of gene expression on leucine starvation in GCN2+ ⁄ + and GCN2) ⁄ ) MEF cells The number of genes exhibiting changes in their expression level after h of leucine starvation Filled bars, expression...

Ngày tải lên: 07/03/2014, 03:20

... distinguish it from ML1p and ML2p Therefore we consider ml3.1p as a gene that encodes an isoform of the protein encoded by ml3p Cloning of the sequences of mlp genes Activity of recombinant A-chains ... product of ml1p variants giving 169 and 246 bp fragments SalI cuts the amplification product of ml2p, giving 66, 121 and 243 bp fragments, and the amlpification product of ml3p and ml3.1p, giving 50 and ... separated on 2% agarose gel and the bands corresponding to genomic fragment (length of 415/430 bp) and competitor fragment (length of 388 bp) were excised Radioactivity of the bands was then measured...

Ngày tải lên: 07/03/2014, 15:20

... Isolation and cloning of the pulchellin A-chain gene fragment Expression, purication and characterization of the recombinant pulchellin A-chain Clones of several RIP-2 toxins, such as ricin and abrin ... association of rPAC and rPBC Fig N-glycosidase activities of rPAC and native pulchellin Yeast ribosomes (20 lg) were incubated with different amounts (10, 5, and 0.1 ng) of rPAC and native pulchellin ... [37]: two negative bands at 222 and 208 nm and a positive peak at 196 nm The CD spectrum shape of refolded rPBC showed one maximum band at 231 nm, two minima at 214 and 206 nm, and a negative to...

Ngày tải lên: 07/03/2014, 16:20

... which contained 20 ng of first strand cDNA, · PCR buffer (20 mM Tris/HCl pH 8.4, 50 mM KCl), 0.2 lM of each primer, 1.5 mM MgCl2 and 0.2 mM of dNTPs PCR Construction and screening of a grass carp genomic ... performed on first-strand cDNAs that were reverse transcribed from total RNA by use of Superscript II reverse transcriptase (Invitrogen) and consisted of 20 ng of first strand cDNA, · PCR buffer ... KCl), lM of each primer, 0.2 mM of dNTPs, 1.5 mM MgCl2 and U of Taq DNA polymerase (Invitrogen) The PCR program consisted of predenaturation at 94 °C for min, followed by 35 cycles of amplification...

Ngày tải lên: 17/03/2014, 03:20

... spike morphology of barley and wheat and in hybrids obtained from their crossing and for modification of the expression of AP2-like genes to modify the spike characteristics of cereals for breeding ... genotypes of H chilense (H1), T aestivum (cv Chinese Spring and a 5Hch addition line of cv Chinese spring), and tritordeum (HT374, HT382 and HT22) wild (H11 and H208) and cultivated barley (H106) and ... (Figure 4B) The expression of wheat and barley AP2-like genes was determined in roots, stems, young leaves, and spikes at various developmental stages, and normalized with the expression of the actin...

Ngày tải lên: 12/08/2014, 03:20

... iM Figure genes bymRNA expression of chemokine and interleukin Relative 9qRT-PCR Relative mRNA expression of chemokine and interleukin genes by qRT-PCR GR-1high and GR-1low PBMo, iMϕ and rAM as ... Figure Validation of chemokine and interleukin genes byqRT-PCR Validation of chemokine and interleukin genes byqRT-PCR PBMo, lung Mϕ and DC were sorted as shown in Fig 1A and 2A mRNA expression was ... indicated by ND tain genes that had not been detected by the array experiments (Table and Fig 4, 5, 6) Isolation and gene expression profiling of subpopulations of PBMo and lung M The microarray...

Ngày tải lên: 12/08/2014, 14:20

... treatment Expression of PRSS11, MTSS1, CLPTM1 and SMYD2 was determined in tumor samples Distribution of samples according to the expression of two trios of genes PRSS11, MTSS1, CLPTM1 and PRSS11, ... distribution of tumor samples according to expression of three genes: (a) PRSS11, MTSS1, CLPTM1 and (b) Three-dimensional distribution of tumor samples according to expression of three genes: (a) ... chemotherapy and offers the advantage of an increased likelihood of breast conservation Many drug regimens have been used for a varied number of cycles and two of the most used drugs, doxorubicin and...

Ngày tải lên: 12/08/2014, 18:22

... egg index, percentage of yolk, percentage of albumen and shell strength [22] DNA isolation, RNA isolation and cDNA synthesis Genomic DNA was extracted by the phenol-chloroform method from blood ... significant at P < 0.05 and are presented as least square means ± standard error Results and discussion Molecular cloning and sequence analysis of duck VLDLR Using RACE and sequence matching techniques, ... carried out with total RNA from heart and a pair of primers flanking the deletion was designed Two bands of 268 and 178 bp were amplified and cloned into T-vector and sequenced, which showed that...

Ngày tải lên: 14/08/2014, 13:21

... Figure Expression analysis of 107 presynaptic genes across mouse tissues and developmental stages Expression analysis of 107 presynaptic genes across mouse tissues and developmental stages Expression ... neuronal-specific expression of presynaptic genes, we first defined a cluster of nine presynaptic genes that were highly and specifically expressed in neuronal tissues Unbiased in vivo and in vitro screens of ... transcripts with high expression in brain tissues and low levels of expression in other tissues (b) Cluster 2: transcripts with moderate expression in brain tissues and low levels of expression in other...

Ngày tải lên: 14/08/2014, 21:20