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  • 99
    Thermo Fisher cdna verso cdna kit
    Expression analysis of mecA in JE2 and mutant strains. <t>RNA</t> was isolated from the exponentially grown (0.5–1.2 OD 600nm ) cultures treated with or without oxicillin (Ox) 4 μg/ml for 60 min in MHB. <t>cDNA</t> was prepared from RNA samples and used as template with appropriate primer sets, and relative expression levels were calculated as mentioned in the Materials and Methods section. Each bar shows the mean and standard deviation of values obtained from three replicates. The effect of mutation and oxacillin on expression of pbp2 and mecA was analyzed by performing multiple pairwise comparisons for pbp2 (uppercase letters) and mecA (lowercase letters) using ANOVA followed by Tukey's post-hoc test, and p -values
    Cdna Verso Cdna Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 312 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    cdna verso cdna kit - by Bioz Stars, 2020-04
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    99
    Millipore cdna synthesis
    Expression analysis of mecA in JE2 and mutant strains. <t>RNA</t> was isolated from the exponentially grown (0.5–1.2 OD 600nm ) cultures treated with or without oxicillin (Ox) 4 μg/ml for 60 min in MHB. <t>cDNA</t> was prepared from RNA samples and used as template with appropriate primer sets, and relative expression levels were calculated as mentioned in the Materials and Methods section. Each bar shows the mean and standard deviation of values obtained from three replicates. The effect of mutation and oxacillin on expression of pbp2 and mecA was analyzed by performing multiple pairwise comparisons for pbp2 (uppercase letters) and mecA (lowercase letters) using ANOVA followed by Tukey's post-hoc test, and p -values
    Cdna Synthesis, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 1822 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Illumina Inc cdnas
    Adapter-tRNA ligation efficiencies with a conventional method and YAMAT-seq. ( A ) Schematic representation of the adapter ligation reactions used in the conventional and YAMAT-seq methods. In the conventional method, tRNA was subjected to 3΄-adapter (3΄-AD) and 5΄-adapter (5΄-AD) ligations catalyzed by truncated Rnl2 and Rnl1, respectively, using the <t>Illumina</t> TruSeq Small RNA Sample Preparation Kit. In contrast, in the YAMAT-seq method, a Y-shaped adapter (Y-AD) is ligated to a tRNA by Rnl2. ( B ) Ligation products of synthetic human cyto tRNA AspGUC and 5΄-AD or 3΄-AD were quantified by real-time qRT-PCR. The amounts of ligation products were determined based on standard curves, and ligation efficiencies were calculated as percentages of ligated tRNA versus input tRNA (set as 100%). Each data set represents the average of three independent experiments with bars showing the SD. ( C ) Ligation products of the indicated native cyto tRNAs in BT-474 total RNA and 5΄-AD or 3΄-AD were quantified by real-time qRT-PCR. Each data set represents the average Ct values of three independent experiments with bars showing the SD. ( D ) Image of 8% native polyacrylamide gel electrophoresis (PAGE) of amplified <t>cDNAs</t> resulting from BT-474 total RNA sequencing by conventional and YAMAT-seq procedures. The total adapter lengths of the conventional and YAMAT-seq procedures are 118 and 140 bp, respectively. The YAMAT-seq-derived cDNAs in the designated Bands 1–3 regions were gel-purified and subjected to cloning; the identified sequences are shown in Supplementary Table S1 . The cDNAs in regions designated with gray lines are expected to contain tRNA fraction.
    Cdnas, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 99/100, based on 2653 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    98
    OriGene cdnas
    Adapter-tRNA ligation efficiencies with a conventional method and YAMAT-seq. ( A ) Schematic representation of the adapter ligation reactions used in the conventional and YAMAT-seq methods. In the conventional method, tRNA was subjected to 3΄-adapter (3΄-AD) and 5΄-adapter (5΄-AD) ligations catalyzed by truncated Rnl2 and Rnl1, respectively, using the <t>Illumina</t> TruSeq Small RNA Sample Preparation Kit. In contrast, in the YAMAT-seq method, a Y-shaped adapter (Y-AD) is ligated to a tRNA by Rnl2. ( B ) Ligation products of synthetic human cyto tRNA AspGUC and 5΄-AD or 3΄-AD were quantified by real-time qRT-PCR. The amounts of ligation products were determined based on standard curves, and ligation efficiencies were calculated as percentages of ligated tRNA versus input tRNA (set as 100%). Each data set represents the average of three independent experiments with bars showing the SD. ( C ) Ligation products of the indicated native cyto tRNAs in BT-474 total RNA and 5΄-AD or 3΄-AD were quantified by real-time qRT-PCR. Each data set represents the average Ct values of three independent experiments with bars showing the SD. ( D ) Image of 8% native polyacrylamide gel electrophoresis (PAGE) of amplified <t>cDNAs</t> resulting from BT-474 total RNA sequencing by conventional and YAMAT-seq procedures. The total adapter lengths of the conventional and YAMAT-seq procedures are 118 and 140 bp, respectively. The YAMAT-seq-derived cDNAs in the designated Bands 1–3 regions were gel-purified and subjected to cloning; the identified sequences are shown in Supplementary Table S1 . The cDNAs in regions designated with gray lines are expected to contain tRNA fraction.
    Cdnas, supplied by OriGene, used in various techniques. Bioz Stars score: 98/100, based on 719 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher cdnas
    Overexpression of caspases is not sufficient to accelerate islet cell apoptosis. 832/13 and primary rat islets were untreated (no virus) or treated with recombinant adenoviruses containing <t>cDNAs</t> corresponding to human Apaf-1, human <t>caspase</t> 3, human caspase 9 or the control gene, β-galactosidase (β-gal), as indicated. 72 h post-infection, 832/13 cells ( A ) and primary rat islets ( B ) were treated with DMSO (control) or thapsigargin (100 nM or 1 μM, respectively) for 18 h. Clarified lysates were examined by immunoblot analysis. Endogenous rat (r) and overexpressed human (h) proteins are labeled accordingly. Of note, Apaf-1 and caspase 9 antibodies only detect human (overexpressed) proteins. ( C , D ) 72 h post adenoviral infection, rat islets were treated with DMSO (control) or thapsigargin (1 μM) for up to 72 h. Islet cells were stained for TUNEL, caspase 3, and insulin and counterstained with DAPI. Cells were imaged using a high content imager and analyzed using Cellomics software. ( C ) Percentages of insulin+ TUNEL+ islet cells are shown. ( D ) Comparison of the percentages of caspase 3+ (overexpressing), TUNEL+ islet cells vs. caspase 3- (non-overexpressing), TUNEL+ islet cells are shown. ( C , D ) Data represent the mean +S.E.M. of 3 independent experiments. * p ≤ 0.05 as compared to DMSO treated cells.
    Cdnas, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 53579 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    87
    Unigene cdnas
    Experimental set up and overall expression profiles. (a) The time course of gene expression was determined during the response of two different fibroblast types; 2091 derived from foreskin and WI-38 derived from fetal lung. Each was treated with either the indicated GFs, epidermal growth factor (EGF), fibroblast growth factor (FGF), platelet derived growth factor (PDGF) or FBS (Serum), and three replicate time courses were run for each treatment. The layout of the samples in the other panels as well as in Figures 2-6 is as shown here. (b) Hierarchical cluster of 1,304 genes with a minimum expression change of twofold in at least 15 array experiments and with data present in at least 80% of all array samples. <t>cDNAs</t> with no known <t>Unigene</t> annotation or mapping to multiple Unigene clusters were removed. Black bars on the right indicate consistently induced genes. (c) Sub-cluster branches containing the consistently induced genes were selected and re-clustered. This set included 237 genes represented by 278 cDNA probes. (d) Consistently repressed genes were selected directly from expression data as described in Materials and methods and clustered. This set included 237 genes represented by 250 cDNA probes.
    Cdnas, supplied by Unigene, used in various techniques. Bioz Stars score: 87/100, based on 346 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Eurofins cdnas
    Experimental set up and overall expression profiles. (a) The time course of gene expression was determined during the response of two different fibroblast types; 2091 derived from foreskin and WI-38 derived from fetal lung. Each was treated with either the indicated GFs, epidermal growth factor (EGF), fibroblast growth factor (FGF), platelet derived growth factor (PDGF) or FBS (Serum), and three replicate time courses were run for each treatment. The layout of the samples in the other panels as well as in Figures 2-6 is as shown here. (b) Hierarchical cluster of 1,304 genes with a minimum expression change of twofold in at least 15 array experiments and with data present in at least 80% of all array samples. <t>cDNAs</t> with no known <t>Unigene</t> annotation or mapping to multiple Unigene clusters were removed. Black bars on the right indicate consistently induced genes. (c) Sub-cluster branches containing the consistently induced genes were selected and re-clustered. This set included 237 genes represented by 278 cDNA probes. (d) Consistently repressed genes were selected directly from expression data as described in Materials and methods and clustered. This set included 237 genes represented by 250 cDNA probes.
    Cdnas, supplied by Eurofins, used in various techniques. Bioz Stars score: 94/100, based on 313 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    98
    GenScript cdnas
    Presence of a C-terminal heterologous tag does not influence the temperature- and polyQ dependent conformational effect. A. Scheme of proteins expressed by constructs encoding exon 1 <t>HTT</t> <t>cDNAs</t> with or without C-terminally fused EGFP. B. TR-FRET values (665 nm/615 nm) obtained at RT or 4°C using 2B7-Tb (1 ng/µl)- MW1-d2 (10 ng/µl) on lysates of HEK293T cells transfected with exon 1 HTT expression constructs (Q16, Q39 or Q72) with or without a C-terminal EGFP moiety, confirming the temperature- and polyQ-dependent conformational effect irrespective of the presence of a C-terminal EGFP fusion. C. Absolute ratios between the maximum value of the TR-FRET signal obtained at 4°C and the maximum value of the TR-FRET signal obtained at RT, referred to the curves in C.
    Cdnas, supplied by GenScript, used in various techniques. Bioz Stars score: 98/100, based on 539 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    TaKaRa cdnas
    Virtual Northern and Western blot analyses demonstrating preferential expression of NF-HEV in HEVECs. A: Virtual Northern blot analysis of NF-HEV expression in HEVECs, PMECs, HUVECs, or placenta tissue. <t>PCR-generated</t> full-length <t>cDNAs</t> from the various types of ECs were electrophoresed on a 1% agarose gel, transferred to nylon filters, and hybridized under high-stringency conditions with a 32 P-labeled human NF-HEV cDNA probe. B: Western blot analysis of extracts of tonsillar stroma, HEVECs, PMECs, or HUVECs with rabbit antibodies to NF-HEV. A single band of ∼30 kd was detected in extracts of tonsillar stroma and HEVECs.
    Cdnas, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 14149 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Addgene inc cdnas
    Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4 , <t>Sox2</t> , Klf4 , and c-Myc <t>cDNAs.</t> Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc . Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4 , SOX2 , KLF4 , and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages. Scale bars, 50 μm.
    Cdnas, supplied by Addgene inc, used in various techniques. Bioz Stars score: 99/100, based on 1067 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    87
    EUROIMMUN cdnas
    Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4 , <t>Sox2</t> , Klf4 , and c-Myc <t>cDNAs.</t> Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc . Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4 , SOX2 , KLF4 , and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages. Scale bars, 50 μm.
    Cdnas, supplied by EUROIMMUN, used in various techniques. Bioz Stars score: 87/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Horizon Discovery cdnas
    Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4 , <t>Sox2</t> , Klf4 , and c-Myc <t>cDNAs.</t> Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc . Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4 , SOX2 , KLF4 , and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages. Scale bars, 50 μm.
    Cdnas, supplied by Horizon Discovery, used in various techniques. Bioz Stars score: 94/100, based on 127 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    SLIT2 LTD cdnas
    Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4 , <t>Sox2</t> , Klf4 , and c-Myc <t>cDNAs.</t> Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc . Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4 , SOX2 , KLF4 , and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages. Scale bars, 50 μm.
    Cdnas, supplied by SLIT2 LTD, used in various techniques. Bioz Stars score: 86/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Operon Biotech cdnas
    Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4 , <t>Sox2</t> , Klf4 , and c-Myc <t>cDNAs.</t> Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc . Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4 , SOX2 , KLF4 , and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages. Scale bars, 50 μm.
    Cdnas, supplied by Operon Biotech, used in various techniques. Bioz Stars score: 85/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Evrogen cdnas
    Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4 , <t>Sox2</t> , Klf4 , and c-Myc <t>cDNAs.</t> Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc . Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4 , SOX2 , KLF4 , and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages. Scale bars, 50 μm.
    Cdnas, supplied by Evrogen, used in various techniques. Bioz Stars score: 94/100, based on 115 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    Bioneer Corporation cdnas
    Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4 , <t>Sox2</t> , Klf4 , and c-Myc <t>cDNAs.</t> Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc . Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4 , SOX2 , KLF4 , and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages. Scale bars, 50 μm.
    Cdnas, supplied by Bioneer Corporation, used in various techniques. Bioz Stars score: 97/100, based on 313 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Meridian Life Science cdnas
    Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4 , <t>Sox2</t> , Klf4 , and c-Myc <t>cDNAs.</t> Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc . Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4 , SOX2 , KLF4 , and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages. Scale bars, 50 μm.
    Cdnas, supplied by Meridian Life Science, used in various techniques. Bioz Stars score: 94/100, based on 299 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Mirus Bio cdnas
    Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4 , <t>Sox2</t> , Klf4 , and c-Myc <t>cDNAs.</t> Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc . Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4 , SOX2 , KLF4 , and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages. Scale bars, 50 μm.
    Cdnas, supplied by Mirus Bio, used in various techniques. Bioz Stars score: 93/100, based on 98 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    R&D Systems cdnas
    Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4 , <t>Sox2</t> , Klf4 , and c-Myc <t>cDNAs.</t> Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc . Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4 , SOX2 , KLF4 , and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages. Scale bars, 50 μm.
    Cdnas, supplied by R&D Systems, used in various techniques. Bioz Stars score: 88/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    87
    Incyte cdnas
    Global analysis of fibroblast gene expression in response to IFN, gB, or HCMV infection. Intensity data from all of the <t>cDNAs</t> on the <t>microarrays</t> obtained at 8 and 24 h were log-transformed, and scatter plots were generated to assess differential gene expression between samples. Pair-wise similarity of gene-expression profiles was measured by the correlation coefficient; r ). The 2-h time points were omitted from this analysis because relatively few genes changed at this early time point.
    Cdnas, supplied by Incyte, used in various techniques. Bioz Stars score: 87/100, based on 56 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Promega cdnas
    Global analysis of fibroblast gene expression in response to IFN, gB, or HCMV infection. Intensity data from all of the <t>cDNAs</t> on the <t>microarrays</t> obtained at 8 and 24 h were log-transformed, and scatter plots were generated to assess differential gene expression between samples. Pair-wise similarity of gene-expression profiles was measured by the correlation coefficient; r ). The 2-h time points were omitted from this analysis because relatively few genes changed at this early time point.
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    Dopamine transporter regulation in the dopaminergic MN9D cell line. (A) MN9D cells were transiently cotransfected with DAT cDNA and either a control shRNA directed against no known mouse gene targets (shRNA-Non-Target) or two different shRNAs directed against the MKP3 coding region (shRNA-B and shRNA-C) and assayed for uptake of dopamine (DA). (B) Naïve (MN9D) and pooled stable MKP3 knock-down MN9D cells used in C and D (MN9D shRNA-C) were immunoblotted for MKP3 and actin expression. (C and D) The same cells were transiently <t>transfected</t> with transporter <t>cDNAs</t> and assayed for uptake of dopamine (DA) or serotonin (5-HT). In all uptake assays (A, C, and D) cells were treated with PMA or vehicle 3–4 d after transfection for 30 min, and uptake assays were carried out for 10 min at room temperature. Results are shown as average ± SEM from at least three individual experiments. Statistically significant differences were calculated using Student's t test (*p
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    Dopamine transporter regulation in the dopaminergic MN9D cell line. (A) MN9D cells were transiently cotransfected with DAT cDNA and either a control shRNA directed against no known mouse gene targets (shRNA-Non-Target) or two different shRNAs directed against the MKP3 coding region (shRNA-B and shRNA-C) and assayed for uptake of dopamine (DA). (B) Naïve (MN9D) and pooled stable MKP3 knock-down MN9D cells used in C and D (MN9D shRNA-C) were immunoblotted for MKP3 and actin expression. (C and D) The same cells were transiently <t>transfected</t> with transporter <t>cDNAs</t> and assayed for uptake of dopamine (DA) or serotonin (5-HT). In all uptake assays (A, C, and D) cells were treated with PMA or vehicle 3–4 d after transfection for 30 min, and uptake assays were carried out for 10 min at room temperature. Results are shown as average ± SEM from at least three individual experiments. Statistically significant differences were calculated using Student's t test (*p
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    Millipore cdna
    Dopamine transporter regulation in the dopaminergic MN9D cell line. (A) MN9D cells were transiently cotransfected with DAT cDNA and either a control shRNA directed against no known mouse gene targets (shRNA-Non-Target) or two different shRNAs directed against the MKP3 coding region (shRNA-B and shRNA-C) and assayed for uptake of dopamine (DA). (B) Naïve (MN9D) and pooled stable MKP3 knock-down MN9D cells used in C and D (MN9D shRNA-C) were immunoblotted for MKP3 and actin expression. (C and D) The same cells were transiently <t>transfected</t> with transporter <t>cDNAs</t> and assayed for uptake of dopamine (DA) or serotonin (5-HT). In all uptake assays (A, C, and D) cells were treated with PMA or vehicle 3–4 d after transfection for 30 min, and uptake assays were carried out for 10 min at room temperature. Results are shown as average ± SEM from at least three individual experiments. Statistically significant differences were calculated using Student's t test (*p
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    Immunohistologic analysis of TELi. (A–B): H E staining of native liver and TELi. An asterisk is placed near a blood vessel containing erythrocytes in TELi. White arrows indicate incompletely degraded scaffold material. (C): Immunofluorescence staining for albumin (red) and CK8 (cholangiocyte and periductular hepatocyte marker, green) in native liver and TELi. Arrowhead shows double positive cells. (D): Immunofluorescence staining for HNF4α (hepatocyte specific transcription factor, red) and CK8 (green). Nuclei stained with 4′,6‐diamidino‐2‐phenylindole (blue). (E): Reverse transcription‐polymerase chain reaction for albumin expression. <t>cDNA</t> prepared from whole liver <t>RNA.</t> Scale bars = 50 μm (A) , 25 μm (B–D) . Data represent more than three independent experiments. Abbreviations: H E, hematoxylin and eosin; L, ladder; NC, negative control (water); PC, positive control; TELi, tissue‐engineered liver.
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    <t>RNA</t> Extraction and <t>cDNA</t> Synthesis
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    TaKaRa cdna ends ready cdna
    Analysis of allelic <t>Commd1</t> expression and Zrsr1 -DMR methylation in PA and WT MII oocytes. a MII oocytes were prepared from Commd1 PA(B6)/ + (PWK) females (PA) and Commd1 + (B6)/ + (PWK) females (WT). RT-PCR was performed as in Fig. 1 b. Amplified <t>cDNA</t> (250 bp) was digested with NlaIII (CATG). There are two NlaIII restriction sites in the B6 amplicon, but one of these is absent in the PWK amplicon because of a single nucleotide polymorphism (SNP) between the two strains. Adult brain RNA from each of the strains was used as the control. MW: molecular weight marker. b A 278-bp region in Zrsr1 -DMR containing 15 CpG sites (B6) or 14 CpG sites (PWK) was analyzed via bisulfite sequencing. The alleles were discriminated via an SNP (C in B6 and A in PWK) located in the leftmost CpG site in the B6 sequence. Closed and open circles depict methylated and unmethylated CpGs, respectively
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    Thermo Fisher cdna
    Improved hierarchical clustering of combined NCI-60 cell-lines profiled by <t>Affymetrix</t> gene-chip and <t>cDNA</t> microarray by sequence-overlapping probe measurements. The gene expression profiles obtained for the sixty cell lines by the Affymetrix gene chips and the Stanford cDNA microarray platform were pooled after data transformation as described in the text. Gene expression data by the two different platforms were matched by either Unigene ID matching or by redefining the Affymetrix probe sets based on the sequence overlap criteria of the probes. The pooled gene expression profiles were subjected to average linkage hierarchical clustering. Matched cell-lines from the two platforms which cluster together are marked by red branches in the dendrogram. ( A ) Unigene-matched measurements tended to cluster the cell-lines by measurement platform, and produced only 28 instances of matched cell-lines clustering together. ( B ) Sequence-overlapping probe measurements produced more (43) instances of matched cell-lines from each platform clustering together.
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    Thermo Fisher primers cdna
    Expression levels of type I collagen, Smad2/3, and TGF-β1 genes after ES-P treatment in aged mice. (A) The left ears of fourteen-week old mice were treated daily with 30 μg T . spiralis ES-P for 14 days (red color). The right ears of were treated daily with PBS for 14 days as control (blue color). After 14 days, ears were collected. Total <t>RNA</t> was extracted from each ear tissue and <t>cDNA</t> was constructed. (B) The gene expression levels of collagen I , TGF-β1 , and Smad2/3 were analyzed by real-time PCR. The gene expression value of each group was normalized by control value, 14 weeks group. (**; P
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    Expression levels of type I collagen, Smad2/3, and TGF-β1 genes after ES-P treatment in aged mice. (A) The left ears of fourteen-week old mice were treated daily with 30 μg T . spiralis ES-P for 14 days (red color). The right ears of were treated daily with PBS for 14 days as control (blue color). After 14 days, ears were collected. Total <t>RNA</t> was extracted from each ear tissue and <t>cDNA</t> was constructed. (B) The gene expression levels of collagen I , TGF-β1 , and Smad2/3 were analyzed by real-time PCR. The gene expression value of each group was normalized by control value, 14 weeks group. (**; P
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    Image Search Results


    Expression analysis of mecA in JE2 and mutant strains. RNA was isolated from the exponentially grown (0.5–1.2 OD 600nm ) cultures treated with or without oxicillin (Ox) 4 μg/ml for 60 min in MHB. cDNA was prepared from RNA samples and used as template with appropriate primer sets, and relative expression levels were calculated as mentioned in the Materials and Methods section. Each bar shows the mean and standard deviation of values obtained from three replicates. The effect of mutation and oxacillin on expression of pbp2 and mecA was analyzed by performing multiple pairwise comparisons for pbp2 (uppercase letters) and mecA (lowercase letters) using ANOVA followed by Tukey's post-hoc test, and p -values

    Journal: Frontiers in Microbiology

    Article Title: RelQ Mediates the Expression of β-Lactam Resistance in Methicillin-Resistant Staphylococcus aureus

    doi: 10.3389/fmicb.2019.00339

    Figure Lengend Snippet: Expression analysis of mecA in JE2 and mutant strains. RNA was isolated from the exponentially grown (0.5–1.2 OD 600nm ) cultures treated with or without oxicillin (Ox) 4 μg/ml for 60 min in MHB. cDNA was prepared from RNA samples and used as template with appropriate primer sets, and relative expression levels were calculated as mentioned in the Materials and Methods section. Each bar shows the mean and standard deviation of values obtained from three replicates. The effect of mutation and oxacillin on expression of pbp2 and mecA was analyzed by performing multiple pairwise comparisons for pbp2 (uppercase letters) and mecA (lowercase letters) using ANOVA followed by Tukey's post-hoc test, and p -values

    Article Snippet: For relative quantification, 2 μg total RNA was reverse transcribed using a cDNA Synthesis Kit (Fermentas, USA), and 1 μl of 10x diluted cDNA was used as template in 20 μl reaction volume.

    Techniques: Expressing, Mutagenesis, Isolation, Standard Deviation

    Effect of relQ expression on mecA transcription in parent and mutant strains. Transcript level was monitored by RT-PCR. cDNA was prepared by reverse-transcription of the RNA samples isolated from exponentially grown cultures treated with or without oxicillin (Ox) 4 μg/ml for 60 min in MHB. The relative expression levels were calculated using the 2 - Δ Δ C T method. Each bar shows the mean and standard deviation of values obtained from three replicates. The effect of relQ expression on mecA transcription level (lowercase letters) in different strains was analyzed by performing multiple pairwise comparisons using ANOVA followed by Tukey's post-hoc test, and p -values

    Journal: Frontiers in Microbiology

    Article Title: RelQ Mediates the Expression of β-Lactam Resistance in Methicillin-Resistant Staphylococcus aureus

    doi: 10.3389/fmicb.2019.00339

    Figure Lengend Snippet: Effect of relQ expression on mecA transcription in parent and mutant strains. Transcript level was monitored by RT-PCR. cDNA was prepared by reverse-transcription of the RNA samples isolated from exponentially grown cultures treated with or without oxicillin (Ox) 4 μg/ml for 60 min in MHB. The relative expression levels were calculated using the 2 - Δ Δ C T method. Each bar shows the mean and standard deviation of values obtained from three replicates. The effect of relQ expression on mecA transcription level (lowercase letters) in different strains was analyzed by performing multiple pairwise comparisons using ANOVA followed by Tukey's post-hoc test, and p -values

    Article Snippet: For relative quantification, 2 μg total RNA was reverse transcribed using a cDNA Synthesis Kit (Fermentas, USA), and 1 μl of 10x diluted cDNA was used as template in 20 μl reaction volume.

    Techniques: Expressing, Mutagenesis, Reverse Transcription Polymerase Chain Reaction, Isolation, Standard Deviation

    mRNA expression of N-extended AChE, AChE-S, and AChE-R isoforms following treatment of Y79 cells with glucose for 1 h. Y79 cells were pre-treated for 16–24 h in starvation medium (1% FBS and 1 mg/ml of glucose) and then with 3.5 mg/ml glucose for different time intervals. Total RNA was extracted and cDNA was prepared for real-time PCR procedure as described under Materials and Methods. Results are presented as fold of control cells cultured in starvation media. Values are means ± SEM, ( N = 4). * p

    Journal: Frontiers in Molecular Neuroscience

    Article Title: Acetylcholinesterase (AChE) is an important link in the apoptotic pathway induced by hyperglycemia in Y79 retinoblastoma cell line

    doi: 10.3389/fnmol.2012.00069

    Figure Lengend Snippet: mRNA expression of N-extended AChE, AChE-S, and AChE-R isoforms following treatment of Y79 cells with glucose for 1 h. Y79 cells were pre-treated for 16–24 h in starvation medium (1% FBS and 1 mg/ml of glucose) and then with 3.5 mg/ml glucose for different time intervals. Total RNA was extracted and cDNA was prepared for real-time PCR procedure as described under Materials and Methods. Results are presented as fold of control cells cultured in starvation media. Values are means ± SEM, ( N = 4). * p

    Article Snippet: RNA samples (0.2 μg) were used for cDNA synthesis prepared by the Verso™ cDNA Kit (Thermo SCIENTIFIC, UK).

    Techniques: Expressing, Real-time Polymerase Chain Reaction, Cell Culture

    Adapter-tRNA ligation efficiencies with a conventional method and YAMAT-seq. ( A ) Schematic representation of the adapter ligation reactions used in the conventional and YAMAT-seq methods. In the conventional method, tRNA was subjected to 3΄-adapter (3΄-AD) and 5΄-adapter (5΄-AD) ligations catalyzed by truncated Rnl2 and Rnl1, respectively, using the Illumina TruSeq Small RNA Sample Preparation Kit. In contrast, in the YAMAT-seq method, a Y-shaped adapter (Y-AD) is ligated to a tRNA by Rnl2. ( B ) Ligation products of synthetic human cyto tRNA AspGUC and 5΄-AD or 3΄-AD were quantified by real-time qRT-PCR. The amounts of ligation products were determined based on standard curves, and ligation efficiencies were calculated as percentages of ligated tRNA versus input tRNA (set as 100%). Each data set represents the average of three independent experiments with bars showing the SD. ( C ) Ligation products of the indicated native cyto tRNAs in BT-474 total RNA and 5΄-AD or 3΄-AD were quantified by real-time qRT-PCR. Each data set represents the average Ct values of three independent experiments with bars showing the SD. ( D ) Image of 8% native polyacrylamide gel electrophoresis (PAGE) of amplified cDNAs resulting from BT-474 total RNA sequencing by conventional and YAMAT-seq procedures. The total adapter lengths of the conventional and YAMAT-seq procedures are 118 and 140 bp, respectively. The YAMAT-seq-derived cDNAs in the designated Bands 1–3 regions were gel-purified and subjected to cloning; the identified sequences are shown in Supplementary Table S1 . The cDNAs in regions designated with gray lines are expected to contain tRNA fraction.

    Journal: Nucleic Acids Research

    Article Title: YAMAT-seq: an efficient method for high-throughput sequencing of mature transfer RNAs

    doi: 10.1093/nar/gkx005

    Figure Lengend Snippet: Adapter-tRNA ligation efficiencies with a conventional method and YAMAT-seq. ( A ) Schematic representation of the adapter ligation reactions used in the conventional and YAMAT-seq methods. In the conventional method, tRNA was subjected to 3΄-adapter (3΄-AD) and 5΄-adapter (5΄-AD) ligations catalyzed by truncated Rnl2 and Rnl1, respectively, using the Illumina TruSeq Small RNA Sample Preparation Kit. In contrast, in the YAMAT-seq method, a Y-shaped adapter (Y-AD) is ligated to a tRNA by Rnl2. ( B ) Ligation products of synthetic human cyto tRNA AspGUC and 5΄-AD or 3΄-AD were quantified by real-time qRT-PCR. The amounts of ligation products were determined based on standard curves, and ligation efficiencies were calculated as percentages of ligated tRNA versus input tRNA (set as 100%). Each data set represents the average of three independent experiments with bars showing the SD. ( C ) Ligation products of the indicated native cyto tRNAs in BT-474 total RNA and 5΄-AD or 3΄-AD were quantified by real-time qRT-PCR. Each data set represents the average Ct values of three independent experiments with bars showing the SD. ( D ) Image of 8% native polyacrylamide gel electrophoresis (PAGE) of amplified cDNAs resulting from BT-474 total RNA sequencing by conventional and YAMAT-seq procedures. The total adapter lengths of the conventional and YAMAT-seq procedures are 118 and 140 bp, respectively. The YAMAT-seq-derived cDNAs in the designated Bands 1–3 regions were gel-purified and subjected to cloning; the identified sequences are shown in Supplementary Table S1 . The cDNAs in regions designated with gray lines are expected to contain tRNA fraction.

    Article Snippet: Fourth, the resultant cDNAs are gel-purified and subjected to Illumina sequencing.

    Techniques: Ligation, Sample Prep, Quantitative RT-PCR, Polyacrylamide Gel Electrophoresis, Amplification, RNA Sequencing Assay, Derivative Assay, Purification, Clone Assay

    Amplification and sequencing of mature tRNAs using YAMAT-seq. ( A ) Image of 8% native PAGE of amplified cDNAs resulting from YAMAT-seq of total RNAs from MCF-7, SK-BR-3 and BT-20 cells. The result of one of the triplicate, Replicate 1 (R1), from each cell line is shown. The region designated with a gray line was subjected to gel-purification and Illumina sequencing. ( B ) Flow diagram of the read numbers filtered by the indicated sequence analyses. The result of R1 from each cell line is shown, whereas the results of R2 and R3 are shown in Supplementary Figure S1 . ( C ) Spearman's rank correlation analysis using tRNA reads of the technical triplicates (R1–R3) from each cell line. The results using the reads of all tRNAs (left), cyto tRNAs (middle) and mt tRNAs (right) are shown. The color bar indicates the strength of Spearman's correlation coefficient.

    Journal: Nucleic Acids Research

    Article Title: YAMAT-seq: an efficient method for high-throughput sequencing of mature transfer RNAs

    doi: 10.1093/nar/gkx005

    Figure Lengend Snippet: Amplification and sequencing of mature tRNAs using YAMAT-seq. ( A ) Image of 8% native PAGE of amplified cDNAs resulting from YAMAT-seq of total RNAs from MCF-7, SK-BR-3 and BT-20 cells. The result of one of the triplicate, Replicate 1 (R1), from each cell line is shown. The region designated with a gray line was subjected to gel-purification and Illumina sequencing. ( B ) Flow diagram of the read numbers filtered by the indicated sequence analyses. The result of R1 from each cell line is shown, whereas the results of R2 and R3 are shown in Supplementary Figure S1 . ( C ) Spearman's rank correlation analysis using tRNA reads of the technical triplicates (R1–R3) from each cell line. The results using the reads of all tRNAs (left), cyto tRNAs (middle) and mt tRNAs (right) are shown. The color bar indicates the strength of Spearman's correlation coefficient.

    Article Snippet: Fourth, the resultant cDNAs are gel-purified and subjected to Illumina sequencing.

    Techniques: Amplification, Sequencing, Clear Native PAGE, Gel Purification, Flow Cytometry

    Overexpression of caspases is not sufficient to accelerate islet cell apoptosis. 832/13 and primary rat islets were untreated (no virus) or treated with recombinant adenoviruses containing cDNAs corresponding to human Apaf-1, human caspase 3, human caspase 9 or the control gene, β-galactosidase (β-gal), as indicated. 72 h post-infection, 832/13 cells ( A ) and primary rat islets ( B ) were treated with DMSO (control) or thapsigargin (100 nM or 1 μM, respectively) for 18 h. Clarified lysates were examined by immunoblot analysis. Endogenous rat (r) and overexpressed human (h) proteins are labeled accordingly. Of note, Apaf-1 and caspase 9 antibodies only detect human (overexpressed) proteins. ( C , D ) 72 h post adenoviral infection, rat islets were treated with DMSO (control) or thapsigargin (1 μM) for up to 72 h. Islet cells were stained for TUNEL, caspase 3, and insulin and counterstained with DAPI. Cells were imaged using a high content imager and analyzed using Cellomics software. ( C ) Percentages of insulin+ TUNEL+ islet cells are shown. ( D ) Comparison of the percentages of caspase 3+ (overexpressing), TUNEL+ islet cells vs. caspase 3- (non-overexpressing), TUNEL+ islet cells are shown. ( C , D ) Data represent the mean +S.E.M. of 3 independent experiments. * p ≤ 0.05 as compared to DMSO treated cells.

    Journal: PLoS ONE

    Article Title: Delayed apoptosis allows islet β-cells to implement an autophagic mechanism to promote cell survival

    doi: 10.1371/journal.pone.0172567

    Figure Lengend Snippet: Overexpression of caspases is not sufficient to accelerate islet cell apoptosis. 832/13 and primary rat islets were untreated (no virus) or treated with recombinant adenoviruses containing cDNAs corresponding to human Apaf-1, human caspase 3, human caspase 9 or the control gene, β-galactosidase (β-gal), as indicated. 72 h post-infection, 832/13 cells ( A ) and primary rat islets ( B ) were treated with DMSO (control) or thapsigargin (100 nM or 1 μM, respectively) for 18 h. Clarified lysates were examined by immunoblot analysis. Endogenous rat (r) and overexpressed human (h) proteins are labeled accordingly. Of note, Apaf-1 and caspase 9 antibodies only detect human (overexpressed) proteins. ( C , D ) 72 h post adenoviral infection, rat islets were treated with DMSO (control) or thapsigargin (1 μM) for up to 72 h. Islet cells were stained for TUNEL, caspase 3, and insulin and counterstained with DAPI. Cells were imaged using a high content imager and analyzed using Cellomics software. ( C ) Percentages of insulin+ TUNEL+ islet cells are shown. ( D ) Comparison of the percentages of caspase 3+ (overexpressing), TUNEL+ islet cells vs. caspase 3- (non-overexpressing), TUNEL+ islet cells are shown. ( C , D ) Data represent the mean +S.E.M. of 3 independent experiments. * p ≤ 0.05 as compared to DMSO treated cells.

    Article Snippet: Plasmids containing cDNAs encoding caspase 3 (HsCD00043626) and caspase 9 (HsCD00043776) were from DNASU [ ]. cDNAs were recombined into pAd/CMV/V5-DEST (Thermo Life) using LR Clonase II plus (Thermo Life) per the manufacturer’s protocol [ ].

    Techniques: Over Expression, Recombinant, Infection, Labeling, Staining, TUNEL Assay, Software

    Experimental set up and overall expression profiles. (a) The time course of gene expression was determined during the response of two different fibroblast types; 2091 derived from foreskin and WI-38 derived from fetal lung. Each was treated with either the indicated GFs, epidermal growth factor (EGF), fibroblast growth factor (FGF), platelet derived growth factor (PDGF) or FBS (Serum), and three replicate time courses were run for each treatment. The layout of the samples in the other panels as well as in Figures 2-6 is as shown here. (b) Hierarchical cluster of 1,304 genes with a minimum expression change of twofold in at least 15 array experiments and with data present in at least 80% of all array samples. cDNAs with no known Unigene annotation or mapping to multiple Unigene clusters were removed. Black bars on the right indicate consistently induced genes. (c) Sub-cluster branches containing the consistently induced genes were selected and re-clustered. This set included 237 genes represented by 278 cDNA probes. (d) Consistently repressed genes were selected directly from expression data as described in Materials and methods and clustered. This set included 237 genes represented by 250 cDNA probes.

    Journal: Genome Biology

    Article Title: PI3K signaling and miRNA expression during the response of quiescent human fibroblasts to distinct proliferative stimuli

    doi: 10.1186/gb-2006-7-5-r42

    Figure Lengend Snippet: Experimental set up and overall expression profiles. (a) The time course of gene expression was determined during the response of two different fibroblast types; 2091 derived from foreskin and WI-38 derived from fetal lung. Each was treated with either the indicated GFs, epidermal growth factor (EGF), fibroblast growth factor (FGF), platelet derived growth factor (PDGF) or FBS (Serum), and three replicate time courses were run for each treatment. The layout of the samples in the other panels as well as in Figures 2-6 is as shown here. (b) Hierarchical cluster of 1,304 genes with a minimum expression change of twofold in at least 15 array experiments and with data present in at least 80% of all array samples. cDNAs with no known Unigene annotation or mapping to multiple Unigene clusters were removed. Black bars on the right indicate consistently induced genes. (c) Sub-cluster branches containing the consistently induced genes were selected and re-clustered. This set included 237 genes represented by 278 cDNA probes. (d) Consistently repressed genes were selected directly from expression data as described in Materials and methods and clustered. This set included 237 genes represented by 250 cDNA probes.

    Article Snippet: After removing cDNAs with no known Unigene annotation or those that matched multiple Unigene clusters, we were left with 1,304 cDNAs that were subjected to hierarchical clustering.

    Techniques: Expressing, Derivative Assay

    Presence of a C-terminal heterologous tag does not influence the temperature- and polyQ dependent conformational effect. A. Scheme of proteins expressed by constructs encoding exon 1 HTT cDNAs with or without C-terminally fused EGFP. B. TR-FRET values (665 nm/615 nm) obtained at RT or 4°C using 2B7-Tb (1 ng/µl)- MW1-d2 (10 ng/µl) on lysates of HEK293T cells transfected with exon 1 HTT expression constructs (Q16, Q39 or Q72) with or without a C-terminal EGFP moiety, confirming the temperature- and polyQ-dependent conformational effect irrespective of the presence of a C-terminal EGFP fusion. C. Absolute ratios between the maximum value of the TR-FRET signal obtained at 4°C and the maximum value of the TR-FRET signal obtained at RT, referred to the curves in C.

    Journal: PLoS ONE

    Article Title: Polyglutamine- and Temperature-Dependent Conformational Rigidity in Mutant Huntingtin Revealed by Immunoassays and Circular Dichroism Spectroscopy

    doi: 10.1371/journal.pone.0112262

    Figure Lengend Snippet: Presence of a C-terminal heterologous tag does not influence the temperature- and polyQ dependent conformational effect. A. Scheme of proteins expressed by constructs encoding exon 1 HTT cDNAs with or without C-terminally fused EGFP. B. TR-FRET values (665 nm/615 nm) obtained at RT or 4°C using 2B7-Tb (1 ng/µl)- MW1-d2 (10 ng/µl) on lysates of HEK293T cells transfected with exon 1 HTT expression constructs (Q16, Q39 or Q72) with or without a C-terminal EGFP moiety, confirming the temperature- and polyQ-dependent conformational effect irrespective of the presence of a C-terminal EGFP fusion. C. Absolute ratios between the maximum value of the TR-FRET signal obtained at 4°C and the maximum value of the TR-FRET signal obtained at RT, referred to the curves in C.

    Article Snippet: Plasmid constructs cDNAs encoding N-terminal fragments (exon 1, N548) or full length human HTT bearing different polyQ lengths were obtained as follows. cDNAs encoding human HTT exon 1 fragments (Q16, Q39 or Q72) were synthesized (Genscript, Piscataway, NJ) and subcloned into pCDNA3.1 either with or without a C-terminal EGFP translational fusion.

    Techniques: Construct, Transfection, Expressing

    Virtual Northern and Western blot analyses demonstrating preferential expression of NF-HEV in HEVECs. A: Virtual Northern blot analysis of NF-HEV expression in HEVECs, PMECs, HUVECs, or placenta tissue. PCR-generated full-length cDNAs from the various types of ECs were electrophoresed on a 1% agarose gel, transferred to nylon filters, and hybridized under high-stringency conditions with a 32 P-labeled human NF-HEV cDNA probe. B: Western blot analysis of extracts of tonsillar stroma, HEVECs, PMECs, or HUVECs with rabbit antibodies to NF-HEV. A single band of ∼30 kd was detected in extracts of tonsillar stroma and HEVECs.

    Journal: The American Journal of Pathology

    Article Title: Molecular Characterization of NF-HEV, a Nuclear Factor Preferentially Expressed in Human High Endothelial Venules

    doi:

    Figure Lengend Snippet: Virtual Northern and Western blot analyses demonstrating preferential expression of NF-HEV in HEVECs. A: Virtual Northern blot analysis of NF-HEV expression in HEVECs, PMECs, HUVECs, or placenta tissue. PCR-generated full-length cDNAs from the various types of ECs were electrophoresed on a 1% agarose gel, transferred to nylon filters, and hybridized under high-stringency conditions with a 32 P-labeled human NF-HEV cDNA probe. B: Western blot analysis of extracts of tonsillar stroma, HEVECs, PMECs, or HUVECs with rabbit antibodies to NF-HEV. A single band of ∼30 kd was detected in extracts of tonsillar stroma and HEVECs.

    Article Snippet: To obtain sufficient amounts of double-stranded (ds) cDNA for subtraction, both PMEC and HEVEC cDNAs were preamplified with the SMART PCR cDNA Synthesis kit (Clontech, Palo Alto, CA). cDNAs synthesized from 1 μg of total RNA (HEVECs) or 0.15 μg mRNA (PMECs) with Advantage KlenTaq polymerase (22 cycles, Clontech) were used with the PCR Select cDNA Subtraction kit (Clontech).

    Techniques: Northern Blot, Western Blot, Expressing, Polymerase Chain Reaction, Generated, Agarose Gel Electrophoresis, Labeling

    Two splice forms of mouse Tgif2 . A) Expression of mouse Tgif2 was analyzed by PCR from 1 st strand cDNA, using primers at the 5' and 3' ends of the coding sequence. cDNAs were from the tissues indicated (sk. musc = skeletal muscle), or from embryos at the indicated days of gestation. -ve: no cDNA. Similar PCR analysis was carried out on cDNA from human brain and kidney (right). B) Alignment of putative splice sequences from mouse and human TGIF2 with splice consensus sequences (matches to the consensus are shaded gray).

    Journal: BMC Molecular Biology

    Article Title: The Tgif2 gene contains a retained intron within the coding sequence

    doi: 10.1186/1471-2199-7-2

    Figure Lengend Snippet: Two splice forms of mouse Tgif2 . A) Expression of mouse Tgif2 was analyzed by PCR from 1 st strand cDNA, using primers at the 5' and 3' ends of the coding sequence. cDNAs were from the tissues indicated (sk. musc = skeletal muscle), or from embryos at the indicated days of gestation. -ve: no cDNA. Similar PCR analysis was carried out on cDNA from human brain and kidney (right). B) Alignment of putative splice sequences from mouse and human TGIF2 with splice consensus sequences (matches to the consensus are shaded gray).

    Article Snippet: Analysis of Tgif2 expression in mouse tissues was carried out by PCR, from first strand cDNAs (Clontech).

    Techniques: Expressing, Polymerase Chain Reaction, Sequencing

    Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc cDNAs. Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc . Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4 , SOX2 , KLF4 , and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages. Scale bars, 50 μm.

    Journal: Stem Cell Reports

    Article Title: NANOG Is Required for the Long-Term Establishment of Avian Somatic Reprogrammed Cells

    doi: 10.1016/j.stemcr.2018.09.005

    Figure Lengend Snippet: Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc cDNAs. Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4 , Sox2 , Klf4 , and c-Myc . Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4 , SOX2 , KLF4 , and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages. Scale bars, 50 μm.

    Article Snippet: The cDNAs from chicken POUV ( POU5F3 ), SOX2 , SOX3 , KLF4 , c-MYC , NANOG , and LIN28 were cloned, sequenced, inserted into the pGAE lentivirus or pPB transposon backbones, and deposited to Addgene.

    Techniques: Infection, Amplification, Isolation

    Global analysis of fibroblast gene expression in response to IFN, gB, or HCMV infection. Intensity data from all of the cDNAs on the microarrays obtained at 8 and 24 h were log-transformed, and scatter plots were generated to assess differential gene expression between samples. Pair-wise similarity of gene-expression profiles was measured by the correlation coefficient; r ). The 2-h time points were omitted from this analysis because relatively few genes changed at this early time point.

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Global modulation of cellular transcription by human cytomegalovirus is initiated by viral glycoprotein B

    doi: 10.1073/pnas.121177598

    Figure Lengend Snippet: Global analysis of fibroblast gene expression in response to IFN, gB, or HCMV infection. Intensity data from all of the cDNAs on the microarrays obtained at 8 and 24 h were log-transformed, and scatter plots were generated to assess differential gene expression between samples. Pair-wise similarity of gene-expression profiles was measured by the correlation coefficient; r ). The 2-h time points were omitted from this analysis because relatively few genes changed at this early time point.

    Article Snippet: The cDNAs printed on the microarrays were from the IMAGE consortium (Integrated Molecular Analysis of Genome and their Expression) and Incyte libraries.

    Techniques: Expressing, Infection, Transformation Assay, Generated

    Dopamine transporter regulation in the dopaminergic MN9D cell line. (A) MN9D cells were transiently cotransfected with DAT cDNA and either a control shRNA directed against no known mouse gene targets (shRNA-Non-Target) or two different shRNAs directed against the MKP3 coding region (shRNA-B and shRNA-C) and assayed for uptake of dopamine (DA). (B) Naïve (MN9D) and pooled stable MKP3 knock-down MN9D cells used in C and D (MN9D shRNA-C) were immunoblotted for MKP3 and actin expression. (C and D) The same cells were transiently transfected with transporter cDNAs and assayed for uptake of dopamine (DA) or serotonin (5-HT). In all uptake assays (A, C, and D) cells were treated with PMA or vehicle 3–4 d after transfection for 30 min, and uptake assays were carried out for 10 min at room temperature. Results are shown as average ± SEM from at least three individual experiments. Statistically significant differences were calculated using Student's t test (*p

    Journal: Molecular Biology of the Cell

    Article Title: Genetic Complementation Screen Identifies a Mitogen-activated Protein Kinase Phosphatase, MKP3, as a Regulator of Dopamine Transporter Trafficking

    doi: 10.1091/mbc.E07-09-0980

    Figure Lengend Snippet: Dopamine transporter regulation in the dopaminergic MN9D cell line. (A) MN9D cells were transiently cotransfected with DAT cDNA and either a control shRNA directed against no known mouse gene targets (shRNA-Non-Target) or two different shRNAs directed against the MKP3 coding region (shRNA-B and shRNA-C) and assayed for uptake of dopamine (DA). (B) Naïve (MN9D) and pooled stable MKP3 knock-down MN9D cells used in C and D (MN9D shRNA-C) were immunoblotted for MKP3 and actin expression. (C and D) The same cells were transiently transfected with transporter cDNAs and assayed for uptake of dopamine (DA) or serotonin (5-HT). In all uptake assays (A, C, and D) cells were treated with PMA or vehicle 3–4 d after transfection for 30 min, and uptake assays were carried out for 10 min at room temperature. Results are shown as average ± SEM from at least three individual experiments. Statistically significant differences were calculated using Student's t test (*p

    Article Snippet: These cells were transfected with cDNAs for transporters using Fugene HD transfection reagent (Roche Applied Science, Indianapolis, IN) according to manufacturer's protocol.

    Techniques: shRNA, Expressing, Transfection

    Immunohistologic analysis of TELi. (A–B): H E staining of native liver and TELi. An asterisk is placed near a blood vessel containing erythrocytes in TELi. White arrows indicate incompletely degraded scaffold material. (C): Immunofluorescence staining for albumin (red) and CK8 (cholangiocyte and periductular hepatocyte marker, green) in native liver and TELi. Arrowhead shows double positive cells. (D): Immunofluorescence staining for HNF4α (hepatocyte specific transcription factor, red) and CK8 (green). Nuclei stained with 4′,6‐diamidino‐2‐phenylindole (blue). (E): Reverse transcription‐polymerase chain reaction for albumin expression. cDNA prepared from whole liver RNA. Scale bars = 50 μm (A) , 25 μm (B–D) . Data represent more than three independent experiments. Abbreviations: H E, hematoxylin and eosin; L, ladder; NC, negative control (water); PC, positive control; TELi, tissue‐engineered liver.

    Journal: Stem Cells Translational Medicine

    Article Title: Functional Human and Murine Tissue‐Engineered Liver Is Generated from Adult Stem/Progenitor Cells

    doi: 10.5966/sctm.2016-0205

    Figure Lengend Snippet: Immunohistologic analysis of TELi. (A–B): H E staining of native liver and TELi. An asterisk is placed near a blood vessel containing erythrocytes in TELi. White arrows indicate incompletely degraded scaffold material. (C): Immunofluorescence staining for albumin (red) and CK8 (cholangiocyte and periductular hepatocyte marker, green) in native liver and TELi. Arrowhead shows double positive cells. (D): Immunofluorescence staining for HNF4α (hepatocyte specific transcription factor, red) and CK8 (green). Nuclei stained with 4′,6‐diamidino‐2‐phenylindole (blue). (E): Reverse transcription‐polymerase chain reaction for albumin expression. cDNA prepared from whole liver RNA. Scale bars = 50 μm (A) , 25 μm (B–D) . Data represent more than three independent experiments. Abbreviations: H E, hematoxylin and eosin; L, ladder; NC, negative control (water); PC, positive control; TELi, tissue‐engineered liver.

    Article Snippet: Reverse Transcription‐Polymerase Chain Reaction RNA was isolated from frozen TELi with the Qiagen RNA purification kit. cDNA was prepared from 1000 ng of RNA using Bio‐Rad cDNA synthesis kit (Bio‐Rad, Hercules, CA, http://www.bio-rad.com ).

    Techniques: Staining, Immunofluorescence, Marker, Reverse Transcription Polymerase Chain Reaction, Expressing, Negative Control, Positive Control

    Breast tumor kinase expression in breast cancer cell lines. (a) Total RNA was isolated from the breast cancer cell lines using the RNeasy kit (Qiagen), and the cDNA was generated using iScript cDNA synthesis (Bio-Rad). The cDNA was amplified using primers specifically for breast tumor kinase (Brk) or β-actin. The relative level of Brk mRNA was determined using densitometric analysis and was normalized to the amount of β-actin. The level of Brk mRNA in the MDA-MB-468 cell line was set to 1; numbers indicate the fold increase compared with the 468 cells. (b) Whole-cell lysates were prepared from the breast cancer cell lines. Lysates were immunoblotted with anti-Brk antibody (top) and with anti-Ran antibody (bottom). The relative expression of Brk protein was determined using densitometric analysis and was normalized to the amount of Ran. The expression of Brk protein in the MDA-MB-468 cell line was set to 1; numbers indicate the fold increase compared with the 468 cells.

    Journal: Breast Cancer Research : BCR

    Article Title: Signal transducer and activator of transcription 5b: a new target of breast tumor kinase/protein tyrosine kinase 6

    doi: 10.1186/bcr1794

    Figure Lengend Snippet: Breast tumor kinase expression in breast cancer cell lines. (a) Total RNA was isolated from the breast cancer cell lines using the RNeasy kit (Qiagen), and the cDNA was generated using iScript cDNA synthesis (Bio-Rad). The cDNA was amplified using primers specifically for breast tumor kinase (Brk) or β-actin. The relative level of Brk mRNA was determined using densitometric analysis and was normalized to the amount of β-actin. The level of Brk mRNA in the MDA-MB-468 cell line was set to 1; numbers indicate the fold increase compared with the 468 cells. (b) Whole-cell lysates were prepared from the breast cancer cell lines. Lysates were immunoblotted with anti-Brk antibody (top) and with anti-Ran antibody (bottom). The relative expression of Brk protein was determined using densitometric analysis and was normalized to the amount of Ran. The expression of Brk protein in the MDA-MB-468 cell line was set to 1; numbers indicate the fold increase compared with the 468 cells.

    Article Snippet: The cDNA was generated using iScript cDNA synthesis (Bio-Rad).

    Techniques: Expressing, Isolation, Generated, Amplification, Multiple Displacement Amplification

    RNA Extraction and cDNA Synthesis

    Journal: Muscle & nerve

    Article Title: GENE AND PROTEIN EXPRESSION ASSOCIATED WITH PROTEIN SYNTHESIS AND BREAKDOWN IN PARAPLEGIC SKELETAL MUSCLE

    doi: 10.1002/mus.20976

    Figure Lengend Snippet: RNA Extraction and cDNA Synthesis

    Article Snippet: Total RNA (1 μg) was reverse transcribed into cDNA according to the manufacturer's directions (iScript; BioRad, Hercules, California).

    Techniques: RNA Extraction

    Analysis of allelic Commd1 expression and Zrsr1 -DMR methylation in PA and WT MII oocytes. a MII oocytes were prepared from Commd1 PA(B6)/ + (PWK) females (PA) and Commd1 + (B6)/ + (PWK) females (WT). RT-PCR was performed as in Fig. 1 b. Amplified cDNA (250 bp) was digested with NlaIII (CATG). There are two NlaIII restriction sites in the B6 amplicon, but one of these is absent in the PWK amplicon because of a single nucleotide polymorphism (SNP) between the two strains. Adult brain RNA from each of the strains was used as the control. MW: molecular weight marker. b A 278-bp region in Zrsr1 -DMR containing 15 CpG sites (B6) or 14 CpG sites (PWK) was analyzed via bisulfite sequencing. The alleles were discriminated via an SNP (C in B6 and A in PWK) located in the leftmost CpG site in the B6 sequence. Closed and open circles depict methylated and unmethylated CpGs, respectively

    Journal: Epigenetics & Chromatin

    Article Title: Growing oocyte-specific transcription-dependent de novo DNA methylation at the imprinted Zrsr1-DMR

    doi: 10.1186/s13072-018-0200-6

    Figure Lengend Snippet: Analysis of allelic Commd1 expression and Zrsr1 -DMR methylation in PA and WT MII oocytes. a MII oocytes were prepared from Commd1 PA(B6)/ + (PWK) females (PA) and Commd1 + (B6)/ + (PWK) females (WT). RT-PCR was performed as in Fig. 1 b. Amplified cDNA (250 bp) was digested with NlaIII (CATG). There are two NlaIII restriction sites in the B6 amplicon, but one of these is absent in the PWK amplicon because of a single nucleotide polymorphism (SNP) between the two strains. Adult brain RNA from each of the strains was used as the control. MW: molecular weight marker. b A 278-bp region in Zrsr1 -DMR containing 15 CpG sites (B6) or 14 CpG sites (PWK) was analyzed via bisulfite sequencing. The alleles were discriminated via an SNP (C in B6 and A in PWK) located in the leftmost CpG site in the B6 sequence. Closed and open circles depict methylated and unmethylated CpGs, respectively

    Article Snippet: Expression analysis To analyze Commd1 expression in oocyte and adult tissues, cDNA was synthesized with random primers (Takara, #3802) and reverse transcriptase (TOYOBO, TRT-101), and the Commd1 cDNA was amplified by PCR with the primers Comm-F1 (exon 1) and Comm-R1 (exon 2).

    Techniques: Expressing, Methylation, Reverse Transcription Polymerase Chain Reaction, Amplification, Molecular Weight, Marker, Methylation Sequencing, Sequencing

    Analysis of Commd1 and Zrsr1 expression and Zrsr1 -DMR methylation in adult mice. a Allelic expression of Commd1 was analyzed as in Fig. 2 a in the brain (Br) and liver (Lv) of Commd1 PA(B6)/ + (PWK) (PA) and Commd1 + (B6)/ + (PWK) (WT) adult F1 mice. MW: molecular weight marker. B6 and PWK are the maternal and paternal alleles in the F1 mice, respectively. b Methylation at Zrsr1 -DMR was analyzed as in Fig. 2 b in the brain of adult F1 mice. Shown is the SNP used to discriminate the parental alleles. c Allelic expression of Zrsr1 was analyzed in the F1 adult brain by direct RT-PCR sequencing of the cDNA, which contains two SNP sites between B6 and PWK alleles. d Total Zrsr1 expression was analyzed in triplicate (shown with n) in the adult F1 brain and liver via TaqMan RT-PCR. Total expression is presented relative to the WT expression level in each tissue. The differences in levels of expression between WT and ΔPA F1 mice were statistically significant in the brain and liver (two-sided Student’s t test, * P

    Journal: Epigenetics & Chromatin

    Article Title: Growing oocyte-specific transcription-dependent de novo DNA methylation at the imprinted Zrsr1-DMR

    doi: 10.1186/s13072-018-0200-6

    Figure Lengend Snippet: Analysis of Commd1 and Zrsr1 expression and Zrsr1 -DMR methylation in adult mice. a Allelic expression of Commd1 was analyzed as in Fig. 2 a in the brain (Br) and liver (Lv) of Commd1 PA(B6)/ + (PWK) (PA) and Commd1 + (B6)/ + (PWK) (WT) adult F1 mice. MW: molecular weight marker. B6 and PWK are the maternal and paternal alleles in the F1 mice, respectively. b Methylation at Zrsr1 -DMR was analyzed as in Fig. 2 b in the brain of adult F1 mice. Shown is the SNP used to discriminate the parental alleles. c Allelic expression of Zrsr1 was analyzed in the F1 adult brain by direct RT-PCR sequencing of the cDNA, which contains two SNP sites between B6 and PWK alleles. d Total Zrsr1 expression was analyzed in triplicate (shown with n) in the adult F1 brain and liver via TaqMan RT-PCR. Total expression is presented relative to the WT expression level in each tissue. The differences in levels of expression between WT and ΔPA F1 mice were statistically significant in the brain and liver (two-sided Student’s t test, * P

    Article Snippet: Expression analysis To analyze Commd1 expression in oocyte and adult tissues, cDNA was synthesized with random primers (Takara, #3802) and reverse transcriptase (TOYOBO, TRT-101), and the Commd1 cDNA was amplified by PCR with the primers Comm-F1 (exon 1) and Comm-R1 (exon 2).

    Techniques: Expressing, Methylation, Mouse Assay, Molecular Weight, Marker, Reverse Transcription Polymerase Chain Reaction, Sequencing

    Structure of the Zrsr1 / Commd1 locus and analysis of Commd1 expression and Zrsr1 -DMR methylation in the oocyte. a Zrsr1 , an approximately 2.8-kb intronless gene, and the first two exons of Commd1 are represented by gray and white boxes, respectively. Distances from the Zrsr1 gene to Commd1 exon 1 and exon 2 are indicated above the gene with double-headed arrows. The schematic is not drawn to scale. Arrows above (maternal allele) and below (paternal allele) exon 1 and the Zrsr1 gene represent the direction of transcription and the allelic expression status of the genes. The open and closed circles at the Zrsr1 promoter indicate unmethylation and methylation, respectively. A schematic of the targeting vector is shown under the gene. The closed and hatched boxes represent the truncation cassette and the neo-selection marker gene, respectively. These elements are flanked by the 5.2 kb left arm containing exon 1 and the 5.1 kb right arm, which contains part of intron 1. The truncation cassette is flanked by loxP sites, represented by gray arrowheads enclosed in open rectangles. Expected transcription patterns of the WT and PA alleles are shown above the gene schematic with thick lines and dotted lines corresponding to exons and introns, respectively. b RT-PCR analysis of Commd1 expression in growing oocytes prepared from B6 female neonates at Day 5 (D5), Day 10 (D10), and Day 15 (D15) postpartum, and fully grown MII oocytes (MII) from B6 adult females. PC: positive control for RT-PCR using adult brain cDNA. MW: molecular weight marker. c Analysis of methylation at Zrsr1 -DMR in growing and fully grown oocytes used in b . The 223-bp region in the DMR containing 14 CpGs was analyzed via bisulfite sequencing. Each row represents a dataset from one clone, and each circle represents one CpG site. Closed and open circles depict methylated and unmethylated CpGs, respectively

    Journal: Epigenetics & Chromatin

    Article Title: Growing oocyte-specific transcription-dependent de novo DNA methylation at the imprinted Zrsr1-DMR

    doi: 10.1186/s13072-018-0200-6

    Figure Lengend Snippet: Structure of the Zrsr1 / Commd1 locus and analysis of Commd1 expression and Zrsr1 -DMR methylation in the oocyte. a Zrsr1 , an approximately 2.8-kb intronless gene, and the first two exons of Commd1 are represented by gray and white boxes, respectively. Distances from the Zrsr1 gene to Commd1 exon 1 and exon 2 are indicated above the gene with double-headed arrows. The schematic is not drawn to scale. Arrows above (maternal allele) and below (paternal allele) exon 1 and the Zrsr1 gene represent the direction of transcription and the allelic expression status of the genes. The open and closed circles at the Zrsr1 promoter indicate unmethylation and methylation, respectively. A schematic of the targeting vector is shown under the gene. The closed and hatched boxes represent the truncation cassette and the neo-selection marker gene, respectively. These elements are flanked by the 5.2 kb left arm containing exon 1 and the 5.1 kb right arm, which contains part of intron 1. The truncation cassette is flanked by loxP sites, represented by gray arrowheads enclosed in open rectangles. Expected transcription patterns of the WT and PA alleles are shown above the gene schematic with thick lines and dotted lines corresponding to exons and introns, respectively. b RT-PCR analysis of Commd1 expression in growing oocytes prepared from B6 female neonates at Day 5 (D5), Day 10 (D10), and Day 15 (D15) postpartum, and fully grown MII oocytes (MII) from B6 adult females. PC: positive control for RT-PCR using adult brain cDNA. MW: molecular weight marker. c Analysis of methylation at Zrsr1 -DMR in growing and fully grown oocytes used in b . The 223-bp region in the DMR containing 14 CpGs was analyzed via bisulfite sequencing. Each row represents a dataset from one clone, and each circle represents one CpG site. Closed and open circles depict methylated and unmethylated CpGs, respectively

    Article Snippet: Expression analysis To analyze Commd1 expression in oocyte and adult tissues, cDNA was synthesized with random primers (Takara, #3802) and reverse transcriptase (TOYOBO, TRT-101), and the Commd1 cDNA was amplified by PCR with the primers Comm-F1 (exon 1) and Comm-R1 (exon 2).

    Techniques: Expressing, Methylation, Plasmid Preparation, Selection, Marker, Reverse Transcription Polymerase Chain Reaction, Positive Control, Molecular Weight, Methylation Sequencing

    The antisense transcript in the promoter region of Commd1 in adult tissues. a Strand-specific RT-PCR was performed to detect an antisense transcript to Commd1 transcription in the promoter region. The arrows represent the primers, CommUP-F1, for cDNA synthesis, and CommUP-F2 and CommUP-R2, for subsequent RT-PCR. The amplicon is 206 bp in size. CommUP-R2 is located approximately 100-bp upstream from the putative transcription start site of Commd1 . b Brain (Br) and liver (Lv) RNA were analyzed. Tissues were prepared from adult F1 mice generated from crossing PA-B6 females with WT PWK males. cDNA synthesis was performed with (RTase +) or without (RTase −) reverse transcriptase. MW: molecular weight marker. c Allelic expression of the antisense transcripts was analyzed via direct sequencing of the amplified brain cDNA from the WT and PA mice shown in b . Three SNPs used to discriminate the parental alleles are shown under the electropherograms

    Journal: Epigenetics & Chromatin

    Article Title: Growing oocyte-specific transcription-dependent de novo DNA methylation at the imprinted Zrsr1-DMR

    doi: 10.1186/s13072-018-0200-6

    Figure Lengend Snippet: The antisense transcript in the promoter region of Commd1 in adult tissues. a Strand-specific RT-PCR was performed to detect an antisense transcript to Commd1 transcription in the promoter region. The arrows represent the primers, CommUP-F1, for cDNA synthesis, and CommUP-F2 and CommUP-R2, for subsequent RT-PCR. The amplicon is 206 bp in size. CommUP-R2 is located approximately 100-bp upstream from the putative transcription start site of Commd1 . b Brain (Br) and liver (Lv) RNA were analyzed. Tissues were prepared from adult F1 mice generated from crossing PA-B6 females with WT PWK males. cDNA synthesis was performed with (RTase +) or without (RTase −) reverse transcriptase. MW: molecular weight marker. c Allelic expression of the antisense transcripts was analyzed via direct sequencing of the amplified brain cDNA from the WT and PA mice shown in b . Three SNPs used to discriminate the parental alleles are shown under the electropherograms

    Article Snippet: Expression analysis To analyze Commd1 expression in oocyte and adult tissues, cDNA was synthesized with random primers (Takara, #3802) and reverse transcriptase (TOYOBO, TRT-101), and the Commd1 cDNA was amplified by PCR with the primers Comm-F1 (exon 1) and Comm-R1 (exon 2).

    Techniques: Reverse Transcription Polymerase Chain Reaction, Amplification, Mouse Assay, Generated, Molecular Weight, Marker, Expressing, Sequencing

    Improved hierarchical clustering of combined NCI-60 cell-lines profiled by Affymetrix gene-chip and cDNA microarray by sequence-overlapping probe measurements. The gene expression profiles obtained for the sixty cell lines by the Affymetrix gene chips and the Stanford cDNA microarray platform were pooled after data transformation as described in the text. Gene expression data by the two different platforms were matched by either Unigene ID matching or by redefining the Affymetrix probe sets based on the sequence overlap criteria of the probes. The pooled gene expression profiles were subjected to average linkage hierarchical clustering. Matched cell-lines from the two platforms which cluster together are marked by red branches in the dendrogram. ( A ) Unigene-matched measurements tended to cluster the cell-lines by measurement platform, and produced only 28 instances of matched cell-lines clustering together. ( B ) Sequence-overlapping probe measurements produced more (43) instances of matched cell-lines from each platform clustering together.

    Journal: BMC Bioinformatics

    Article Title: Redefinition of Affymetrix probe sets by sequence overlap with cDNA microarray probes reduces cross-platform inconsistencies in cancer-associated gene expression measurements

    doi: 10.1186/1471-2105-6-107

    Figure Lengend Snippet: Improved hierarchical clustering of combined NCI-60 cell-lines profiled by Affymetrix gene-chip and cDNA microarray by sequence-overlapping probe measurements. The gene expression profiles obtained for the sixty cell lines by the Affymetrix gene chips and the Stanford cDNA microarray platform were pooled after data transformation as described in the text. Gene expression data by the two different platforms were matched by either Unigene ID matching or by redefining the Affymetrix probe sets based on the sequence overlap criteria of the probes. The pooled gene expression profiles were subjected to average linkage hierarchical clustering. Matched cell-lines from the two platforms which cluster together are marked by red branches in the dendrogram. ( A ) Unigene-matched measurements tended to cluster the cell-lines by measurement platform, and produced only 28 instances of matched cell-lines clustering together. ( B ) Sequence-overlapping probe measurements produced more (43) instances of matched cell-lines from each platform clustering together.

    Article Snippet: However, for a clear presentation of results we chose to compare the following classes representing different levels of shared identity: a) Affymetrix probe sets that share a Unigene ID with a cDNA clone. (termed Shared Unigene probes) b) Affymetrix probe sets containing probes that could be sequence-matched to the same transcript sequence as the cDNA clone, but for which no Affymetrix probe actually overlaps the cDNA clone sequence (termed Shared Transcript probes); c) Affymetrix probe sets with 1 to 10 probes sequence overlapping with the cDNA clone (termed Partially Overlapping probes); d) Affymetrix probe sets with 20 (i.e. all) probes sequence overlapping with the cDNA clone (termed Completely Overlapping probes); e) alt-CDF or "redefined probe sets" for which all probes across the entire array that matched to a given cDNA clone insert were used to define a new derivative probe set.

    Techniques: Chromatin Immunoprecipitation, Microarray, Sequencing, Expressing, Transformation Assay, Produced

    Increased efficiency of breast cancer subtype classification transfer from cDNA microarray to Affymetrix HG-U95Av2 gene-chip tumor-profiles by sequence-overlapping probe measurements. Tumor samples profiled on the Affymetrix platform were classified according to their correlation with the set of subtype median-centroids derived from cDNA microarray measurements (see methods). The classified samples were then hierarchically clustered using Pearson correlation and average-linkage agglomeration. ( A ), Affymetrix measurements matched to the cDNA centroids by Unigene identifier. ( B ), Affymetrix measurements matched to cDNA centroids by sequence-overlap of probe features produced more coherent classifications. In particular, the large ERbB2+ subtype cluster (upper left) is mostly absent from the unigene-based classification. The significance of this cluster is supported by the observation that all tumors in this cluster for which Her-2 amplification was assessed by immunohistochemistry were designated positive.

    Journal: BMC Bioinformatics

    Article Title: Redefinition of Affymetrix probe sets by sequence overlap with cDNA microarray probes reduces cross-platform inconsistencies in cancer-associated gene expression measurements

    doi: 10.1186/1471-2105-6-107

    Figure Lengend Snippet: Increased efficiency of breast cancer subtype classification transfer from cDNA microarray to Affymetrix HG-U95Av2 gene-chip tumor-profiles by sequence-overlapping probe measurements. Tumor samples profiled on the Affymetrix platform were classified according to their correlation with the set of subtype median-centroids derived from cDNA microarray measurements (see methods). The classified samples were then hierarchically clustered using Pearson correlation and average-linkage agglomeration. ( A ), Affymetrix measurements matched to the cDNA centroids by Unigene identifier. ( B ), Affymetrix measurements matched to cDNA centroids by sequence-overlap of probe features produced more coherent classifications. In particular, the large ERbB2+ subtype cluster (upper left) is mostly absent from the unigene-based classification. The significance of this cluster is supported by the observation that all tumors in this cluster for which Her-2 amplification was assessed by immunohistochemistry were designated positive.

    Article Snippet: However, for a clear presentation of results we chose to compare the following classes representing different levels of shared identity: a) Affymetrix probe sets that share a Unigene ID with a cDNA clone. (termed Shared Unigene probes) b) Affymetrix probe sets containing probes that could be sequence-matched to the same transcript sequence as the cDNA clone, but for which no Affymetrix probe actually overlaps the cDNA clone sequence (termed Shared Transcript probes); c) Affymetrix probe sets with 1 to 10 probes sequence overlapping with the cDNA clone (termed Partially Overlapping probes); d) Affymetrix probe sets with 20 (i.e. all) probes sequence overlapping with the cDNA clone (termed Completely Overlapping probes); e) alt-CDF or "redefined probe sets" for which all probes across the entire array that matched to a given cDNA clone insert were used to define a new derivative probe set.

    Techniques: Microarray, Chromatin Immunoprecipitation, Sequencing, Derivative Assay, Produced, Amplification, Immunohistochemistry

    Sequence-overlapping probes give greater cross-platform concordance for the NCI-60 panel. ( A ) Pearson correlation coefficient was calculated for each gene between its expression values measured on the Affymetrix Hu6800 platforms and its expression values measured on the Stanford cDNA microarray across sixty cell lines of the NCI-60 panel. The figure shows the cumulative distribution of the Pearson correlation coefficients for all genes analyzed. The five different curves reflect the level of cross-platform consistency of probe sets with various levels of overlap between the two microarray platforms. Matched gene measurements across the two platforms showed higher correlation when greater numbers of probes in the Affymetrix probe sets overlapped the insert region of the cDNA clone. The highest correlation was attained when only those Affymetrix probes overlapping the insert-sequence of a given cDNA clone were retained. Measurements for which the probes targeted the same transcript as the cDNA clone, but did not overlap the clone sequence, showed the lowest correlation. ( B ), Pearson correlation coefficient was calculated across all genes for each matched sample pair profiled by the Affymetrix Hu6800 platform and by the Stanford cDNA microarray. The figure shows the cumulative distribution of the Pearson correlation coefficients for the sixty cell lines of the NCI-60 panel. Matched cell-line measurements showed identical stratification of correlation levels by feature-matching criteria.

    Journal: BMC Bioinformatics

    Article Title: Redefinition of Affymetrix probe sets by sequence overlap with cDNA microarray probes reduces cross-platform inconsistencies in cancer-associated gene expression measurements

    doi: 10.1186/1471-2105-6-107

    Figure Lengend Snippet: Sequence-overlapping probes give greater cross-platform concordance for the NCI-60 panel. ( A ) Pearson correlation coefficient was calculated for each gene between its expression values measured on the Affymetrix Hu6800 platforms and its expression values measured on the Stanford cDNA microarray across sixty cell lines of the NCI-60 panel. The figure shows the cumulative distribution of the Pearson correlation coefficients for all genes analyzed. The five different curves reflect the level of cross-platform consistency of probe sets with various levels of overlap between the two microarray platforms. Matched gene measurements across the two platforms showed higher correlation when greater numbers of probes in the Affymetrix probe sets overlapped the insert region of the cDNA clone. The highest correlation was attained when only those Affymetrix probes overlapping the insert-sequence of a given cDNA clone were retained. Measurements for which the probes targeted the same transcript as the cDNA clone, but did not overlap the clone sequence, showed the lowest correlation. ( B ), Pearson correlation coefficient was calculated across all genes for each matched sample pair profiled by the Affymetrix Hu6800 platform and by the Stanford cDNA microarray. The figure shows the cumulative distribution of the Pearson correlation coefficients for the sixty cell lines of the NCI-60 panel. Matched cell-line measurements showed identical stratification of correlation levels by feature-matching criteria.

    Article Snippet: However, for a clear presentation of results we chose to compare the following classes representing different levels of shared identity: a) Affymetrix probe sets that share a Unigene ID with a cDNA clone. (termed Shared Unigene probes) b) Affymetrix probe sets containing probes that could be sequence-matched to the same transcript sequence as the cDNA clone, but for which no Affymetrix probe actually overlaps the cDNA clone sequence (termed Shared Transcript probes); c) Affymetrix probe sets with 1 to 10 probes sequence overlapping with the cDNA clone (termed Partially Overlapping probes); d) Affymetrix probe sets with 20 (i.e. all) probes sequence overlapping with the cDNA clone (termed Completely Overlapping probes); e) alt-CDF or "redefined probe sets" for which all probes across the entire array that matched to a given cDNA clone insert were used to define a new derivative probe set.

    Techniques: Sequencing, Expressing, Microarray

    Conserved clustering pattern of the NCI-60 cell lines profiled using cDNA microarray and Affymetrix gene chips. Data was normalized as described (methods). Average linkage Pearson correlation hierarchical clustering was computed for each dataset. Cell line names are colored according to cancer type.

    Journal: BMC Bioinformatics

    Article Title: Redefinition of Affymetrix probe sets by sequence overlap with cDNA microarray probes reduces cross-platform inconsistencies in cancer-associated gene expression measurements

    doi: 10.1186/1471-2105-6-107

    Figure Lengend Snippet: Conserved clustering pattern of the NCI-60 cell lines profiled using cDNA microarray and Affymetrix gene chips. Data was normalized as described (methods). Average linkage Pearson correlation hierarchical clustering was computed for each dataset. Cell line names are colored according to cancer type.

    Article Snippet: However, for a clear presentation of results we chose to compare the following classes representing different levels of shared identity: a) Affymetrix probe sets that share a Unigene ID with a cDNA clone. (termed Shared Unigene probes) b) Affymetrix probe sets containing probes that could be sequence-matched to the same transcript sequence as the cDNA clone, but for which no Affymetrix probe actually overlaps the cDNA clone sequence (termed Shared Transcript probes); c) Affymetrix probe sets with 1 to 10 probes sequence overlapping with the cDNA clone (termed Partially Overlapping probes); d) Affymetrix probe sets with 20 (i.e. all) probes sequence overlapping with the cDNA clone (termed Completely Overlapping probes); e) alt-CDF or "redefined probe sets" for which all probes across the entire array that matched to a given cDNA clone insert were used to define a new derivative probe set.

    Techniques: Microarray

    Increased efficiency of breast cancer subtype classification transfer from cDNA microarray to Affymetrix HuFL gene-chip tumor-profiles by sequence-overlapping probe measurements. Tumor samples profiled on the Affymetrix platform were classified according to their correlation with the set of subtype median-centroids derived from cDNA microarray measurements (see methods). The classified samples were then hierarchically clustered using Pearson correlation and average-linkage agglomeration. Affymetrix measurements matched to cDNA centroids by sequence-overlap of probe features produced more coherent classifications than those obtained in the original transfer (Sørlie), specifically, more coherent Luminal A and ERBB2+ subtype clusters.

    Journal: BMC Bioinformatics

    Article Title: Redefinition of Affymetrix probe sets by sequence overlap with cDNA microarray probes reduces cross-platform inconsistencies in cancer-associated gene expression measurements

    doi: 10.1186/1471-2105-6-107

    Figure Lengend Snippet: Increased efficiency of breast cancer subtype classification transfer from cDNA microarray to Affymetrix HuFL gene-chip tumor-profiles by sequence-overlapping probe measurements. Tumor samples profiled on the Affymetrix platform were classified according to their correlation with the set of subtype median-centroids derived from cDNA microarray measurements (see methods). The classified samples were then hierarchically clustered using Pearson correlation and average-linkage agglomeration. Affymetrix measurements matched to cDNA centroids by sequence-overlap of probe features produced more coherent classifications than those obtained in the original transfer (Sørlie), specifically, more coherent Luminal A and ERBB2+ subtype clusters.

    Article Snippet: However, for a clear presentation of results we chose to compare the following classes representing different levels of shared identity: a) Affymetrix probe sets that share a Unigene ID with a cDNA clone. (termed Shared Unigene probes) b) Affymetrix probe sets containing probes that could be sequence-matched to the same transcript sequence as the cDNA clone, but for which no Affymetrix probe actually overlaps the cDNA clone sequence (termed Shared Transcript probes); c) Affymetrix probe sets with 1 to 10 probes sequence overlapping with the cDNA clone (termed Partially Overlapping probes); d) Affymetrix probe sets with 20 (i.e. all) probes sequence overlapping with the cDNA clone (termed Completely Overlapping probes); e) alt-CDF or "redefined probe sets" for which all probes across the entire array that matched to a given cDNA clone insert were used to define a new derivative probe set.

    Techniques: Microarray, Chromatin Immunoprecipitation, Sequencing, Derivative Assay, Produced

    Composition of redefined Affymetrix probe-sets based on overlap with cDNA clone insert sequence. Stacked histograms show the distribution of probe-set size for sets consisting of a single Affymetrix-defined probe-set (black) and for those comprised of probes originally grouped into separate probe-sets by Affymetrix (gray). A , NCI-60 10 k cDNA microarray to HuFL alternative CDF. B , Breast cancer 8 k cDNA microarray to HuFL alternative CDF. C , Breast cancer 8 k cDNA microarray to HG-U95Av2 alternative CDF. D , Lung cancer 22 k cDNA microarray to HG-U95Av2 alternative CDF.

    Journal: BMC Bioinformatics

    Article Title: Redefinition of Affymetrix probe sets by sequence overlap with cDNA microarray probes reduces cross-platform inconsistencies in cancer-associated gene expression measurements

    doi: 10.1186/1471-2105-6-107

    Figure Lengend Snippet: Composition of redefined Affymetrix probe-sets based on overlap with cDNA clone insert sequence. Stacked histograms show the distribution of probe-set size for sets consisting of a single Affymetrix-defined probe-set (black) and for those comprised of probes originally grouped into separate probe-sets by Affymetrix (gray). A , NCI-60 10 k cDNA microarray to HuFL alternative CDF. B , Breast cancer 8 k cDNA microarray to HuFL alternative CDF. C , Breast cancer 8 k cDNA microarray to HG-U95Av2 alternative CDF. D , Lung cancer 22 k cDNA microarray to HG-U95Av2 alternative CDF.

    Article Snippet: However, for a clear presentation of results we chose to compare the following classes representing different levels of shared identity: a) Affymetrix probe sets that share a Unigene ID with a cDNA clone. (termed Shared Unigene probes) b) Affymetrix probe sets containing probes that could be sequence-matched to the same transcript sequence as the cDNA clone, but for which no Affymetrix probe actually overlaps the cDNA clone sequence (termed Shared Transcript probes); c) Affymetrix probe sets with 1 to 10 probes sequence overlapping with the cDNA clone (termed Partially Overlapping probes); d) Affymetrix probe sets with 20 (i.e. all) probes sequence overlapping with the cDNA clone (termed Completely Overlapping probes); e) alt-CDF or "redefined probe sets" for which all probes across the entire array that matched to a given cDNA clone insert were used to define a new derivative probe set.

    Techniques: Sequencing, Microarray

    Expression levels of type I collagen, Smad2/3, and TGF-β1 genes after ES-P treatment in aged mice. (A) The left ears of fourteen-week old mice were treated daily with 30 μg T . spiralis ES-P for 14 days (red color). The right ears of were treated daily with PBS for 14 days as control (blue color). After 14 days, ears were collected. Total RNA was extracted from each ear tissue and cDNA was constructed. (B) The gene expression levels of collagen I , TGF-β1 , and Smad2/3 were analyzed by real-time PCR. The gene expression value of each group was normalized by control value, 14 weeks group. (**; P

    Journal: PLoS Neglected Tropical Diseases

    Article Title: Identification of a host collagen inducing factor from the excretory secretory proteins of Trichinella spiralis

    doi: 10.1371/journal.pntd.0006516

    Figure Lengend Snippet: Expression levels of type I collagen, Smad2/3, and TGF-β1 genes after ES-P treatment in aged mice. (A) The left ears of fourteen-week old mice were treated daily with 30 μg T . spiralis ES-P for 14 days (red color). The right ears of were treated daily with PBS for 14 days as control (blue color). After 14 days, ears were collected. Total RNA was extracted from each ear tissue and cDNA was constructed. (B) The gene expression levels of collagen I , TGF-β1 , and Smad2/3 were analyzed by real-time PCR. The gene expression value of each group was normalized by control value, 14 weeks group. (**; P

    Article Snippet: Real-time PCR Homogenized ear tissues were mixed with TRIzol (Invitrogen, Germany), and total RNA extraction and cDNA synthesis (Invitrogen, Germany) was performed in accordance with the manufacturer’s protocols.

    Techniques: Expressing, Mouse Assay, Construct, Real-time Polymerase Chain Reaction

    Effect of OPN on the expression of genes corresponding to autoimmune and inflammatory responses. ( A ) The analyses were performed using an autoimmune and inflammatory response cDNA array system. A representative experiment is shown as dot blots representing

    Journal: Journal of Clinical Investigation

    Article Title: Role of osteopontin in amplification and perpetuation of rheumatoid synovitis

    doi: 10.1172/JCI200523273

    Figure Lengend Snippet: Effect of OPN on the expression of genes corresponding to autoimmune and inflammatory responses. ( A ) The analyses were performed using an autoimmune and inflammatory response cDNA array system. A representative experiment is shown as dot blots representing

    Article Snippet: Random hexamers were used to prime cDNA synthesis. mRNA expression of OPN, the OPN receptors, and the selected chemokines was determined by real-time PCR using SYBR Green Master Mix (Applied Biosystems).

    Techniques: Expressing