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  • 88
    Promega complementary dna cdna synthesis kit
    A, Ethidium bromide-stained agarose gel of dsRNA. Lanes a and b contain dsRNA extracts from two representative strains of Lentinula edodes FMRI0339 <t>mycovirus</t> (LeV-FMRI0339) and Pleurotus ostreatus ASI2792 mycovirus (ASI2792-PoV) with the characteristic 12-kb viral genome of LeV and approximately 8.0-kb viral genome of ASI2792-PoV, respectively. Lanes 1 to 14 contain dsRNA extracts from 14 isolates obtained by the mycelia fragmentation method described in “Materials and Methods”; B, Northern blot analysis of the corresponding gel with the dsRNA using a partial <t>cDNA</t> clone (792 bp) encoding the RNA-dependent RNA polymerase (RdRp) protein in the PoV-ASI2792 as a probe. Arrowheads indicate the mycovirus genome segment from PoV-ASI2792.
    Complementary Dna Cdna Synthesis Kit, supplied by Promega, used in various techniques. Bioz Stars score: 88/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Bioneer Corporation complementary dna cdna synthesis kit
    A, Ethidium bromide-stained agarose gel of dsRNA. Lanes a and b contain dsRNA extracts from two representative strains of Lentinula edodes FMRI0339 <t>mycovirus</t> (LeV-FMRI0339) and Pleurotus ostreatus ASI2792 mycovirus (ASI2792-PoV) with the characteristic 12-kb viral genome of LeV and approximately 8.0-kb viral genome of ASI2792-PoV, respectively. Lanes 1 to 14 contain dsRNA extracts from 14 isolates obtained by the mycelia fragmentation method described in “Materials and Methods”; B, Northern blot analysis of the corresponding gel with the dsRNA using a partial <t>cDNA</t> clone (792 bp) encoding the RNA-dependent RNA polymerase (RdRp) protein in the PoV-ASI2792 as a probe. Arrowheads indicate the mycovirus genome segment from PoV-ASI2792.
    Complementary Dna Cdna Synthesis Kit, supplied by Bioneer Corporation, used in various techniques. Bioz Stars score: 88/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher complementary dna cdna synthesis kit
    The Total <t>RNA</t> Detection system. A universal RNA adaptor was firstly added to the 3′ ends of all RNA templates. The abundant 18S and 28S ribosome RNAs were then subtracted by using biotinylated ribosomal-specific probes. Small-size RNAs containing the degraded RNA intermediates were removed by size-filtration. The enriched transcripts were converted into double-strand <t>cDNA</t> by using the 3′ end RNA adaptor-based primer. The cDNAs were further digested by NlaIII. The 3′ cDNAs were isolated by using the streptoavidin beads. An adaptor was added to the 5′ ends of the 3′ cDNAs. The 3′ cDNAs were then amplified by PCR using the 5′ adaptor-based sense primer and the 3′ end RNA adaptor-based antisense primer. The amplified 3′ cDNAs were sequenced from the 3′ end by the 454 system. See further details in Materials and Methods .
    Complementary Dna Cdna Synthesis Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 88/100, based on 66 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa complementary dna cdna synthesis kit
    Effects of DCA on mRNA and protein levels of OCTN2 in cancer cell lines. (A), analysis of mRNA levels of OCTN2 in DCA-treated cancer cell lines. After treatment with 0.5 and 1 μM DCA, total <t>RNA</t> in cancer cells was purified by TRIzol reagent for <t>cDNA</t> synthesis and the mRNA levels of OCTN2 were analyzed by Quantitative RT-PCR. (B), analysis of protein levels of OCTN2 in DCA-treated cancer cell lines. Total proteins were extracted and protein levels of OCTN2 were analyzed by western blotting. The relative expression of mRNA and protein was normalized to β-actin. Data presented represent the mean ± S.D. of three independent experiments. Significant difference from control group of 0.1% DMSO is denoted with asterisks (*, p
    Complementary Dna Cdna Synthesis Kit, supplied by TaKaRa, used in various techniques. Bioz Stars score: 90/100, based on 23 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Bio-Rad complementary dna cdna synthesis kit
    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.
    Complementary Dna Cdna Synthesis Kit, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 99/100, based on 24 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Quanta Biosciences cdna synthesis kit
    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.
    Cdna Synthesis Kit, supplied by Quanta Biosciences, used in various techniques. Bioz Stars score: 93/100, based on 332 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Roche cdna synthesis kit
    Messenger <t>RNA</t> expression of TNC receptors of MCF-7 cells after TGF-β and TNC treatments. ( a ) mRNA was extracted from cells co-stimulated with TNC (10 μg/ml) and TGF-β1 (5 ng/ml) for synthesis of <t>cDNA</t> and performance of real-time PCR determining the cycle threshold ( C t ) values for each subunit. Differences in C t values of TNC receptor mRNAs from that of GAPDH mRNA in TGF-β1/TNC-treated cells are shown using qPCR. C t values of α7/8/9 and β3 integrin subunits were very high, indicating only limited expression. Calibration curves were obtained using each target gene in the samples. ( b ) Changes of receptor mRNA expression for αv/2 and β1/6 after treatment with TGF-β1 and/or TNC. The results were normalized to GAPDH expression. Integrin αv showed significant increase after TGF-β1/TNC treatment and β6 was dramatically increased after TGF-β1 addition, with or without TNC (mean±s.d.; * P
    Cdna Synthesis Kit, supplied by Roche, used in various techniques. Bioz Stars score: 93/100, based on 1388 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Quanta Biosciences superscript complementary dna cdna synthesis kit
    Messenger <t>RNA</t> expression of TNC receptors of MCF-7 cells after TGF-β and TNC treatments. ( a ) mRNA was extracted from cells co-stimulated with TNC (10 μg/ml) and TGF-β1 (5 ng/ml) for synthesis of <t>cDNA</t> and performance of real-time PCR determining the cycle threshold ( C t ) values for each subunit. Differences in C t values of TNC receptor mRNAs from that of GAPDH mRNA in TGF-β1/TNC-treated cells are shown using qPCR. C t values of α7/8/9 and β3 integrin subunits were very high, indicating only limited expression. Calibration curves were obtained using each target gene in the samples. ( b ) Changes of receptor mRNA expression for αv/2 and β1/6 after treatment with TGF-β1 and/or TNC. The results were normalized to GAPDH expression. Integrin αv showed significant increase after TGF-β1/TNC treatment and β6 was dramatically increased after TGF-β1 addition, with or without TNC (mean±s.d.; * P
    Superscript Complementary Dna Cdna Synthesis Kit, supplied by Quanta Biosciences, used in various techniques. Bioz Stars score: 93/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TransGen biotech co cdna synthesis kit
    Messenger <t>RNA</t> expression of TNC receptors of MCF-7 cells after TGF-β and TNC treatments. ( a ) mRNA was extracted from cells co-stimulated with TNC (10 μg/ml) and TGF-β1 (5 ng/ml) for synthesis of <t>cDNA</t> and performance of real-time PCR determining the cycle threshold ( C t ) values for each subunit. Differences in C t values of TNC receptor mRNAs from that of GAPDH mRNA in TGF-β1/TNC-treated cells are shown using qPCR. C t values of α7/8/9 and β3 integrin subunits were very high, indicating only limited expression. Calibration curves were obtained using each target gene in the samples. ( b ) Changes of receptor mRNA expression for αv/2 and β1/6 after treatment with TGF-β1 and/or TNC. The results were normalized to GAPDH expression. Integrin αv showed significant increase after TGF-β1/TNC treatment and β6 was dramatically increased after TGF-β1 addition, with or without TNC (mean±s.d.; * P
    Cdna Synthesis Kit, supplied by TransGen biotech co, used in various techniques. Bioz Stars score: 93/100, based on 179 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher complementary dna cdna synthesis supermix kit
    Messenger <t>RNA</t> expression of TNC receptors of MCF-7 cells after TGF-β and TNC treatments. ( a ) mRNA was extracted from cells co-stimulated with TNC (10 μg/ml) and TGF-β1 (5 ng/ml) for synthesis of <t>cDNA</t> and performance of real-time PCR determining the cycle threshold ( C t ) values for each subunit. Differences in C t values of TNC receptor mRNAs from that of GAPDH mRNA in TGF-β1/TNC-treated cells are shown using qPCR. C t values of α7/8/9 and β3 integrin subunits were very high, indicating only limited expression. Calibration curves were obtained using each target gene in the samples. ( b ) Changes of receptor mRNA expression for αv/2 and β1/6 after treatment with TGF-β1 and/or TNC. The results were normalized to GAPDH expression. Integrin αv showed significant increase after TGF-β1/TNC treatment and β6 was dramatically increased after TGF-β1 addition, with or without TNC (mean±s.d.; * P
    Complementary Dna Cdna Synthesis Supermix Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Stratagene cdna synthesis kit
    The nucleotide and deduced amino acid sequences of Fus p 4.0101 (GenBank accession no. KF151224). Numbers to the right are the positions of the nucleotides and the deduced residues of the sequences. The stop codon TAA is denoted with an asterisk (*). The potential N-glycosylation site is indicated in bold letters and boxed. Eight conserved amino acid residues that are involved in the catalysis and the substrate binding of transaldolases are shaded and in bold letters. Nucleotide sequences in grey correspond to primer sequences synthesized for <t>PCR</t> experiments in the <t>cDNA</t> cloning of Fus p 4.0101 as described in the Materials and Methods. The sequences corresponding to primers Fu-TAase-f and Fu-TAase-r used in the preparation of rFus p 4.0101 are boxed.
    Cdna Synthesis Kit, supplied by Stratagene, used in various techniques. Bioz Stars score: 92/100, based on 1002 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Agilent technologies cdna synthesis kit
    Identification of Oplr16 as a pluripotent lncRNA. ( A ) Identification of Oplr16 as a pluripotency-associated lncRNA by integrating RNA-seq and CRIST-seq. RNA-seq was used to identify RNAs that are differentially expressed between fibroblasts and iPSCs collected in the process of reprogramming. In CRIST-seq, the Oct4 promoter was targeted by the expression of the Cas9 Oct4 -gRNA. LncRNAs interacting with Oct4 promoter were reverse transcribed in situ into <t>cDNA</t> with biotin-dCTP. After immunoprecipitation, biotin-cDNAs were purified by streptavidin beads for <t>Illumina</t> sequencing. Both sets of databases were integrated to identify the pluripotency-associated lncRNA Oplr16 . ( B ) Reactivation of Oplr16 in reprogramming. Fibroblasts were transfected with a Oct4-Sox2-Kilf4-c-Myc (OSKM) lentivirus. Cells were collected at various stages of reprogramming and expression of Oplr16 was measured by qPCR. E14: mouse embryonic pluripotent stem cell line used as a positive control; iPSC: induced pluripotent stem cells; non-iPSC: un-reprogrammed cells that express four OSKM factors, but fail to complete reprogramming; FBC: fibroblasts. β-Actin was used as the PCR control. ** P
    Cdna Synthesis Kit, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 93/100, based on 732 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Meridian Life Science cdna synthesis kit
    Identification of Oplr16 as a pluripotent lncRNA. ( A ) Identification of Oplr16 as a pluripotency-associated lncRNA by integrating RNA-seq and CRIST-seq. RNA-seq was used to identify RNAs that are differentially expressed between fibroblasts and iPSCs collected in the process of reprogramming. In CRIST-seq, the Oct4 promoter was targeted by the expression of the Cas9 Oct4 -gRNA. LncRNAs interacting with Oct4 promoter were reverse transcribed in situ into <t>cDNA</t> with biotin-dCTP. After immunoprecipitation, biotin-cDNAs were purified by streptavidin beads for <t>Illumina</t> sequencing. Both sets of databases were integrated to identify the pluripotency-associated lncRNA Oplr16 . ( B ) Reactivation of Oplr16 in reprogramming. Fibroblasts were transfected with a Oct4-Sox2-Kilf4-c-Myc (OSKM) lentivirus. Cells were collected at various stages of reprogramming and expression of Oplr16 was measured by qPCR. E14: mouse embryonic pluripotent stem cell line used as a positive control; iPSC: induced pluripotent stem cells; non-iPSC: un-reprogrammed cells that express four OSKM factors, but fail to complete reprogramming; FBC: fibroblasts. β-Actin was used as the PCR control. ** P
    Cdna Synthesis Kit, supplied by Meridian Life Science, used in various techniques. Bioz Stars score: 92/100, based on 482 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Promega goscript complementary dna cdna synthesis kit
    Identification of Oplr16 as a pluripotent lncRNA. ( A ) Identification of Oplr16 as a pluripotency-associated lncRNA by integrating RNA-seq and CRIST-seq. RNA-seq was used to identify RNAs that are differentially expressed between fibroblasts and iPSCs collected in the process of reprogramming. In CRIST-seq, the Oct4 promoter was targeted by the expression of the Cas9 Oct4 -gRNA. LncRNAs interacting with Oct4 promoter were reverse transcribed in situ into <t>cDNA</t> with biotin-dCTP. After immunoprecipitation, biotin-cDNAs were purified by streptavidin beads for <t>Illumina</t> sequencing. Both sets of databases were integrated to identify the pluripotency-associated lncRNA Oplr16 . ( B ) Reactivation of Oplr16 in reprogramming. Fibroblasts were transfected with a Oct4-Sox2-Kilf4-c-Myc (OSKM) lentivirus. Cells were collected at various stages of reprogramming and expression of Oplr16 was measured by qPCR. E14: mouse embryonic pluripotent stem cell line used as a positive control; iPSC: induced pluripotent stem cells; non-iPSC: un-reprogrammed cells that express four OSKM factors, but fail to complete reprogramming; FBC: fibroblasts. β-Actin was used as the PCR control. ** P
    Goscript Complementary Dna Cdna Synthesis Kit, supplied by Promega, used in various techniques. Bioz Stars score: 92/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Qiagen complementary dna cdna synthesis kit
    Identification of Oplr16 as a pluripotent lncRNA. ( A ) Identification of Oplr16 as a pluripotency-associated lncRNA by integrating RNA-seq and CRIST-seq. RNA-seq was used to identify RNAs that are differentially expressed between fibroblasts and iPSCs collected in the process of reprogramming. In CRIST-seq, the Oct4 promoter was targeted by the expression of the Cas9 Oct4 -gRNA. LncRNAs interacting with Oct4 promoter were reverse transcribed in situ into <t>cDNA</t> with biotin-dCTP. After immunoprecipitation, biotin-cDNAs were purified by streptavidin beads for <t>Illumina</t> sequencing. Both sets of databases were integrated to identify the pluripotency-associated lncRNA Oplr16 . ( B ) Reactivation of Oplr16 in reprogramming. Fibroblasts were transfected with a Oct4-Sox2-Kilf4-c-Myc (OSKM) lentivirus. Cells were collected at various stages of reprogramming and expression of Oplr16 was measured by qPCR. E14: mouse embryonic pluripotent stem cell line used as a positive control; iPSC: induced pluripotent stem cells; non-iPSC: un-reprogrammed cells that express four OSKM factors, but fail to complete reprogramming; FBC: fibroblasts. β-Actin was used as the PCR control. ** P
    Complementary Dna Cdna Synthesis Kit, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Roche first strand complementary dna cdna synthesis kit
    Flow chart for comprehensive NF1 mutation detection. Point mutations identified by <t>DNA</t> sequencing with specific primers (step 1) represented 68.8% of the NF1 mutations. Frameshift and nonsense mutations were identified in 31.3% and 18.8% of NF1 patients, respectively. In addition, missense and splice-site mutations were confirmed using <t>cDNA</t> sequencing (step 2) and were observed in 12.5% and 6.3% of NF1 patients, respectively. In the case of a negative result using DNA sequencing, an analysis of NF1 complete and large partial deletions was performed using multiplex ligation-dependent probe amplification (MLPA) (step 3) and occurred in 25.0% of NF1 patients. This comprehensive mutation screening procedure enabled us to identify an NF1 mutation in 93.8% of the NF1 patients in our study. NF1 = neurofibromatosis type 1.
    First Strand Complementary Dna Cdna Synthesis Kit, supplied by Roche, used in various techniques. Bioz Stars score: 88/100, based on 22 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher complementary dna cdna
    Flow chart for comprehensive NF1 mutation detection. Point mutations identified by <t>DNA</t> sequencing with specific primers (step 1) represented 68.8% of the NF1 mutations. Frameshift and nonsense mutations were identified in 31.3% and 18.8% of NF1 patients, respectively. In addition, missense and splice-site mutations were confirmed using <t>cDNA</t> sequencing (step 2) and were observed in 12.5% and 6.3% of NF1 patients, respectively. In the case of a negative result using DNA sequencing, an analysis of NF1 complete and large partial deletions was performed using multiplex ligation-dependent probe amplification (MLPA) (step 3) and occurred in 25.0% of NF1 patients. This comprehensive mutation screening procedure enabled us to identify an NF1 mutation in 93.8% of the NF1 patients in our study. NF1 = neurofibromatosis type 1.
    Complementary Dna Cdna, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 26028 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Quanta Biosciences qscript cdna synthesis kit
    Flow chart for comprehensive NF1 mutation detection. Point mutations identified by <t>DNA</t> sequencing with specific primers (step 1) represented 68.8% of the NF1 mutations. Frameshift and nonsense mutations were identified in 31.3% and 18.8% of NF1 patients, respectively. In addition, missense and splice-site mutations were confirmed using <t>cDNA</t> sequencing (step 2) and were observed in 12.5% and 6.3% of NF1 patients, respectively. In the case of a negative result using DNA sequencing, an analysis of NF1 complete and large partial deletions was performed using multiplex ligation-dependent probe amplification (MLPA) (step 3) and occurred in 25.0% of NF1 patients. This comprehensive mutation screening procedure enabled us to identify an NF1 mutation in 93.8% of the NF1 patients in our study. NF1 = neurofibromatosis type 1.
    Qscript Cdna Synthesis Kit, supplied by Quanta Biosciences, used in various techniques. Bioz Stars score: 94/100, based on 4783 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    A, Ethidium bromide-stained agarose gel of dsRNA. Lanes a and b contain dsRNA extracts from two representative strains of Lentinula edodes FMRI0339 mycovirus (LeV-FMRI0339) and Pleurotus ostreatus ASI2792 mycovirus (ASI2792-PoV) with the characteristic 12-kb viral genome of LeV and approximately 8.0-kb viral genome of ASI2792-PoV, respectively. Lanes 1 to 14 contain dsRNA extracts from 14 isolates obtained by the mycelia fragmentation method described in “Materials and Methods”; B, Northern blot analysis of the corresponding gel with the dsRNA using a partial cDNA clone (792 bp) encoding the RNA-dependent RNA polymerase (RdRp) protein in the PoV-ASI2792 as a probe. Arrowheads indicate the mycovirus genome segment from PoV-ASI2792.

    Journal: Mycobiology

    Article Title: Viral Effects of a dsRNA Mycovirus (PoV-ASI2792) on the Vegetative Growth of the Edible Mushroom Pleurotus ostreatus

    doi: 10.5941/MYCO.2016.44.4.283

    Figure Lengend Snippet: A, Ethidium bromide-stained agarose gel of dsRNA. Lanes a and b contain dsRNA extracts from two representative strains of Lentinula edodes FMRI0339 mycovirus (LeV-FMRI0339) and Pleurotus ostreatus ASI2792 mycovirus (ASI2792-PoV) with the characteristic 12-kb viral genome of LeV and approximately 8.0-kb viral genome of ASI2792-PoV, respectively. Lanes 1 to 14 contain dsRNA extracts from 14 isolates obtained by the mycelia fragmentation method described in “Materials and Methods”; B, Northern blot analysis of the corresponding gel with the dsRNA using a partial cDNA clone (792 bp) encoding the RNA-dependent RNA polymerase (RdRp) protein in the PoV-ASI2792 as a probe. Arrowheads indicate the mycovirus genome segment from PoV-ASI2792.

    Article Snippet: Cloning and sequence analysis of the partial RdRp gene To obtain a cDNA clone that corresponded to the mycovirus RdRp, cDNA synthesis was conducted as described procedure using a cDNA synthesis kit (Promega, Fitchburg, WI, USA) and reverse transcription-PCR (RT-PCR) was performed as described previously [ ].

    Techniques: Staining, Agarose Gel Electrophoresis, Northern Blot

    The Total RNA Detection system. A universal RNA adaptor was firstly added to the 3′ ends of all RNA templates. The abundant 18S and 28S ribosome RNAs were then subtracted by using biotinylated ribosomal-specific probes. Small-size RNAs containing the degraded RNA intermediates were removed by size-filtration. The enriched transcripts were converted into double-strand cDNA by using the 3′ end RNA adaptor-based primer. The cDNAs were further digested by NlaIII. The 3′ cDNAs were isolated by using the streptoavidin beads. An adaptor was added to the 5′ ends of the 3′ cDNAs. The 3′ cDNAs were then amplified by PCR using the 5′ adaptor-based sense primer and the 3′ end RNA adaptor-based antisense primer. The amplified 3′ cDNAs were sequenced from the 3′ end by the 454 system. See further details in Materials and Methods .

    Journal: PLoS ONE

    Article Title: Poly A- Transcripts Expressed in HeLa Cells

    doi: 10.1371/journal.pone.0002803

    Figure Lengend Snippet: The Total RNA Detection system. A universal RNA adaptor was firstly added to the 3′ ends of all RNA templates. The abundant 18S and 28S ribosome RNAs were then subtracted by using biotinylated ribosomal-specific probes. Small-size RNAs containing the degraded RNA intermediates were removed by size-filtration. The enriched transcripts were converted into double-strand cDNA by using the 3′ end RNA adaptor-based primer. The cDNAs were further digested by NlaIII. The 3′ cDNAs were isolated by using the streptoavidin beads. An adaptor was added to the 5′ ends of the 3′ cDNAs. The 3′ cDNAs were then amplified by PCR using the 5′ adaptor-based sense primer and the 3′ end RNA adaptor-based antisense primer. The amplified 3′ cDNAs were sequenced from the 3′ end by the 454 system. See further details in Materials and Methods .

    Article Snippet: cDNA synthesis and 3′ ESTs collection The enriched RNA was converted into double-strand cDNA by using a cDNA synthesis kit (Invitrogen) following the manufacturer's protocol, except for using the biotin-labeled primer based on the 3′ end adaptor sequences for the priming ( 5′ biotin-ATCTAGAGCGGCCGCAATGGCCACTCTGCGTTGATAC ).

    Techniques: RNA Detection, Filtration, Isolation, Amplification, Polymerase Chain Reaction

    Effects of DCA on mRNA and protein levels of OCTN2 in cancer cell lines. (A), analysis of mRNA levels of OCTN2 in DCA-treated cancer cell lines. After treatment with 0.5 and 1 μM DCA, total RNA in cancer cells was purified by TRIzol reagent for cDNA synthesis and the mRNA levels of OCTN2 were analyzed by Quantitative RT-PCR. (B), analysis of protein levels of OCTN2 in DCA-treated cancer cell lines. Total proteins were extracted and protein levels of OCTN2 were analyzed by western blotting. The relative expression of mRNA and protein was normalized to β-actin. Data presented represent the mean ± S.D. of three independent experiments. Significant difference from control group of 0.1% DMSO is denoted with asterisks (*, p

    Journal: PLoS ONE

    Article Title: Different Involvement of Promoter Methylation in the Expression of Organic Cation/Carnitine Transporter 2 (OCTN2) in Cancer Cell Lines

    doi: 10.1371/journal.pone.0076474

    Figure Lengend Snippet: Effects of DCA on mRNA and protein levels of OCTN2 in cancer cell lines. (A), analysis of mRNA levels of OCTN2 in DCA-treated cancer cell lines. After treatment with 0.5 and 1 μM DCA, total RNA in cancer cells was purified by TRIzol reagent for cDNA synthesis and the mRNA levels of OCTN2 were analyzed by Quantitative RT-PCR. (B), analysis of protein levels of OCTN2 in DCA-treated cancer cell lines. Total proteins were extracted and protein levels of OCTN2 were analyzed by western blotting. The relative expression of mRNA and protein was normalized to β-actin. Data presented represent the mean ± S.D. of three independent experiments. Significant difference from control group of 0.1% DMSO is denoted with asterisks (*, p

    Article Snippet: RNA purification, cDNA Synthesis and Quantitative Real-time PCR Total cellular RNA was isolated from the treated cells using TRIzol reagent. cDNA was synthesized from 1 μg of total RNA using the first cDNA synthesis kit (Takara, Dalian).

    Techniques: Purification, Quantitative RT-PCR, Western Blot, Expressing

    Expression of OCTN2 in cancer cell lines. (A), analysis of mRNA levels of OCTN2 in cancer cell lines. Total RNA in cancer cells was purified by TRIzol reagent for cDNA synthesis and the mRNA levels of OCTN2 were analyzed by Quantitative RT-PCR. (B), analysis of protein levels of OCTN2 in cancer cells. Total proteins were extracted and protein levels of OCTN2 were analyzed by western blotting. The relative expression of mRNA and protein was normalized to β-actin. The expression of HepG2 was set as 1. All of mRNA and protein analysis were performed three times. Significant difference from HepG2 or LS174T cell is denoted with asterisk (*, p

    Journal: PLoS ONE

    Article Title: Different Involvement of Promoter Methylation in the Expression of Organic Cation/Carnitine Transporter 2 (OCTN2) in Cancer Cell Lines

    doi: 10.1371/journal.pone.0076474

    Figure Lengend Snippet: Expression of OCTN2 in cancer cell lines. (A), analysis of mRNA levels of OCTN2 in cancer cell lines. Total RNA in cancer cells was purified by TRIzol reagent for cDNA synthesis and the mRNA levels of OCTN2 were analyzed by Quantitative RT-PCR. (B), analysis of protein levels of OCTN2 in cancer cells. Total proteins were extracted and protein levels of OCTN2 were analyzed by western blotting. The relative expression of mRNA and protein was normalized to β-actin. The expression of HepG2 was set as 1. All of mRNA and protein analysis were performed three times. Significant difference from HepG2 or LS174T cell is denoted with asterisk (*, p

    Article Snippet: RNA purification, cDNA Synthesis and Quantitative Real-time PCR Total cellular RNA was isolated from the treated cells using TRIzol reagent. cDNA was synthesized from 1 μg of total RNA using the first cDNA synthesis kit (Takara, Dalian).

    Techniques: Expressing, Purification, Quantitative RT-PCR, Western Blot

    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

    Messenger RNA expression of TNC receptors of MCF-7 cells after TGF-β and TNC treatments. ( a ) mRNA was extracted from cells co-stimulated with TNC (10 μg/ml) and TGF-β1 (5 ng/ml) for synthesis of cDNA and performance of real-time PCR determining the cycle threshold ( C t ) values for each subunit. Differences in C t values of TNC receptor mRNAs from that of GAPDH mRNA in TGF-β1/TNC-treated cells are shown using qPCR. C t values of α7/8/9 and β3 integrin subunits were very high, indicating only limited expression. Calibration curves were obtained using each target gene in the samples. ( b ) Changes of receptor mRNA expression for αv/2 and β1/6 after treatment with TGF-β1 and/or TNC. The results were normalized to GAPDH expression. Integrin αv showed significant increase after TGF-β1/TNC treatment and β6 was dramatically increased after TGF-β1 addition, with or without TNC (mean±s.d.; * P

    Journal: Oncogenesis

    Article Title: Binding of ?v?1 and ?v?6 integrins to tenascin-C induces epithelial-mesenchymal transition-like change of breast cancer cells

    doi: 10.1038/oncsis.2013.27

    Figure Lengend Snippet: Messenger RNA expression of TNC receptors of MCF-7 cells after TGF-β and TNC treatments. ( a ) mRNA was extracted from cells co-stimulated with TNC (10 μg/ml) and TGF-β1 (5 ng/ml) for synthesis of cDNA and performance of real-time PCR determining the cycle threshold ( C t ) values for each subunit. Differences in C t values of TNC receptor mRNAs from that of GAPDH mRNA in TGF-β1/TNC-treated cells are shown using qPCR. C t values of α7/8/9 and β3 integrin subunits were very high, indicating only limited expression. Calibration curves were obtained using each target gene in the samples. ( b ) Changes of receptor mRNA expression for αv/2 and β1/6 after treatment with TGF-β1 and/or TNC. The results were normalized to GAPDH expression. Integrin αv showed significant increase after TGF-β1/TNC treatment and β6 was dramatically increased after TGF-β1 addition, with or without TNC (mean±s.d.; * P

    Article Snippet: Total RNA (1 μg) was reverse-transcribed using a cDNA synthesis kit (Roche Diagnostics) with anchored-oligo (dT)18 primers.

    Techniques: RNA Expression, Real-time Polymerase Chain Reaction, Expressing

    The nucleotide and deduced amino acid sequences of Fus p 4.0101 (GenBank accession no. KF151224). Numbers to the right are the positions of the nucleotides and the deduced residues of the sequences. The stop codon TAA is denoted with an asterisk (*). The potential N-glycosylation site is indicated in bold letters and boxed. Eight conserved amino acid residues that are involved in the catalysis and the substrate binding of transaldolases are shaded and in bold letters. Nucleotide sequences in grey correspond to primer sequences synthesized for PCR experiments in the cDNA cloning of Fus p 4.0101 as described in the Materials and Methods. The sequences corresponding to primers Fu-TAase-f and Fu-TAase-r used in the preparation of rFus p 4.0101 are boxed.

    Journal: PLoS ONE

    Article Title: The Transaldolase, a Novel Allergen of Fusarium proliferatum, Demonstrates IgE Cross-Reactivity with Its Human Analogue

    doi: 10.1371/journal.pone.0103488

    Figure Lengend Snippet: The nucleotide and deduced amino acid sequences of Fus p 4.0101 (GenBank accession no. KF151224). Numbers to the right are the positions of the nucleotides and the deduced residues of the sequences. The stop codon TAA is denoted with an asterisk (*). The potential N-glycosylation site is indicated in bold letters and boxed. Eight conserved amino acid residues that are involved in the catalysis and the substrate binding of transaldolases are shaded and in bold letters. Nucleotide sequences in grey correspond to primer sequences synthesized for PCR experiments in the cDNA cloning of Fus p 4.0101 as described in the Materials and Methods. The sequences corresponding to primers Fu-TAase-f and Fu-TAase-r used in the preparation of rFus p 4.0101 are boxed.

    Article Snippet: cDNA Cloning The cDNA encoding the F. proliferatum transaldolase was isolated with polymerase chain reactions (PCR) using an AffinityScript Multiple Temperature cDNA Synthesis kit (Stratagene, La Jolla, Calif., USA) as previously described , .

    Techniques: Binding Assay, Synthesized, Polymerase Chain Reaction, Clone Assay

    Identification of Oplr16 as a pluripotent lncRNA. ( A ) Identification of Oplr16 as a pluripotency-associated lncRNA by integrating RNA-seq and CRIST-seq. RNA-seq was used to identify RNAs that are differentially expressed between fibroblasts and iPSCs collected in the process of reprogramming. In CRIST-seq, the Oct4 promoter was targeted by the expression of the Cas9 Oct4 -gRNA. LncRNAs interacting with Oct4 promoter were reverse transcribed in situ into cDNA with biotin-dCTP. After immunoprecipitation, biotin-cDNAs were purified by streptavidin beads for Illumina sequencing. Both sets of databases were integrated to identify the pluripotency-associated lncRNA Oplr16 . ( B ) Reactivation of Oplr16 in reprogramming. Fibroblasts were transfected with a Oct4-Sox2-Kilf4-c-Myc (OSKM) lentivirus. Cells were collected at various stages of reprogramming and expression of Oplr16 was measured by qPCR. E14: mouse embryonic pluripotent stem cell line used as a positive control; iPSC: induced pluripotent stem cells; non-iPSC: un-reprogrammed cells that express four OSKM factors, but fail to complete reprogramming; FBC: fibroblasts. β-Actin was used as the PCR control. ** P

    Journal: Nucleic Acids Research

    Article Title: Oplr16 serves as a novel chromatin factor to control stem cell fate by modulating pluripotency-specific chromosomal looping and TET2-mediated DNA demethylation

    doi: 10.1093/nar/gkaa097

    Figure Lengend Snippet: Identification of Oplr16 as a pluripotent lncRNA. ( A ) Identification of Oplr16 as a pluripotency-associated lncRNA by integrating RNA-seq and CRIST-seq. RNA-seq was used to identify RNAs that are differentially expressed between fibroblasts and iPSCs collected in the process of reprogramming. In CRIST-seq, the Oct4 promoter was targeted by the expression of the Cas9 Oct4 -gRNA. LncRNAs interacting with Oct4 promoter were reverse transcribed in situ into cDNA with biotin-dCTP. After immunoprecipitation, biotin-cDNAs were purified by streptavidin beads for Illumina sequencing. Both sets of databases were integrated to identify the pluripotency-associated lncRNA Oplr16 . ( B ) Reactivation of Oplr16 in reprogramming. Fibroblasts were transfected with a Oct4-Sox2-Kilf4-c-Myc (OSKM) lentivirus. Cells were collected at various stages of reprogramming and expression of Oplr16 was measured by qPCR. E14: mouse embryonic pluripotent stem cell line used as a positive control; iPSC: induced pluripotent stem cells; non-iPSC: un-reprogrammed cells that express four OSKM factors, but fail to complete reprogramming; FBC: fibroblasts. β-Actin was used as the PCR control. ** P

    Article Snippet: The second strand cDNA was synthesized by using Stratagene cDNA Synthesis kit (Agilent Technologies, CA, USA) for Illumina lnc-cDNA sequencing.

    Techniques: RNA Sequencing Assay, Expressing, In Situ, Immunoprecipitation, Purification, Sequencing, Transfection, Real-time Polymerase Chain Reaction, Positive Control, Polymerase Chain Reaction

    Oplr16 binds to the Oct4 promoter. ( A ) Schematic diagram of the RNA reverse transcription-associated trap sequencing (RAT-seq) assay. Oplr16 lncRNA was in situ reverse transcribed using three Oplr16 -specific complementary primers with biotin-dCTP. The random primers were used as the negative control (RAT-CT). After nuclear lysis, the bitoin- Oplr16 cDNA chromatin complex was isolated by streptavidin beads and the Oplr16 -interacting target DNAs were isolated for Illumina library sequencing. ( B ) The IGV analysis of Oplr16 binding signals in the Oct4 locus. 5′-Enh, 3′-Enh: the Oct4 5′- and 3′-enhancers; pOct4: Oct4 promoter; E1-E5: Oct4 exons. ( C ) Quantitative PCR mapping of Oplr16 binding in the Oct4 locus. The RAT pulldown complex was used to map the Oplr16 binding. 5′-CT, 3′-CT: the RAT control sites in the Oct4 locus. Note the enrichment of the Oplr16 binding signals in the promoter region (Prot-1 and Prot-2) ( N = 9, ** P

    Journal: Nucleic Acids Research

    Article Title: Oplr16 serves as a novel chromatin factor to control stem cell fate by modulating pluripotency-specific chromosomal looping and TET2-mediated DNA demethylation

    doi: 10.1093/nar/gkaa097

    Figure Lengend Snippet: Oplr16 binds to the Oct4 promoter. ( A ) Schematic diagram of the RNA reverse transcription-associated trap sequencing (RAT-seq) assay. Oplr16 lncRNA was in situ reverse transcribed using three Oplr16 -specific complementary primers with biotin-dCTP. The random primers were used as the negative control (RAT-CT). After nuclear lysis, the bitoin- Oplr16 cDNA chromatin complex was isolated by streptavidin beads and the Oplr16 -interacting target DNAs were isolated for Illumina library sequencing. ( B ) The IGV analysis of Oplr16 binding signals in the Oct4 locus. 5′-Enh, 3′-Enh: the Oct4 5′- and 3′-enhancers; pOct4: Oct4 promoter; E1-E5: Oct4 exons. ( C ) Quantitative PCR mapping of Oplr16 binding in the Oct4 locus. The RAT pulldown complex was used to map the Oplr16 binding. 5′-CT, 3′-CT: the RAT control sites in the Oct4 locus. Note the enrichment of the Oplr16 binding signals in the promoter region (Prot-1 and Prot-2) ( N = 9, ** P

    Article Snippet: The second strand cDNA was synthesized by using Stratagene cDNA Synthesis kit (Agilent Technologies, CA, USA) for Illumina lnc-cDNA sequencing.

    Techniques: Sequencing, In Situ, Negative Control, Lysis, Isolation, Binding Assay, Real-time Polymerase Chain Reaction

    Flow chart for comprehensive NF1 mutation detection. Point mutations identified by DNA sequencing with specific primers (step 1) represented 68.8% of the NF1 mutations. Frameshift and nonsense mutations were identified in 31.3% and 18.8% of NF1 patients, respectively. In addition, missense and splice-site mutations were confirmed using cDNA sequencing (step 2) and were observed in 12.5% and 6.3% of NF1 patients, respectively. In the case of a negative result using DNA sequencing, an analysis of NF1 complete and large partial deletions was performed using multiplex ligation-dependent probe amplification (MLPA) (step 3) and occurred in 25.0% of NF1 patients. This comprehensive mutation screening procedure enabled us to identify an NF1 mutation in 93.8% of the NF1 patients in our study. NF1 = neurofibromatosis type 1.

    Journal: Medicine

    Article Title: Clinical and Molecular Characterization of NF1 Patients

    doi: 10.1097/MD.0000000000003043

    Figure Lengend Snippet: Flow chart for comprehensive NF1 mutation detection. Point mutations identified by DNA sequencing with specific primers (step 1) represented 68.8% of the NF1 mutations. Frameshift and nonsense mutations were identified in 31.3% and 18.8% of NF1 patients, respectively. In addition, missense and splice-site mutations were confirmed using cDNA sequencing (step 2) and were observed in 12.5% and 6.3% of NF1 patients, respectively. In the case of a negative result using DNA sequencing, an analysis of NF1 complete and large partial deletions was performed using multiplex ligation-dependent probe amplification (MLPA) (step 3) and occurred in 25.0% of NF1 patients. This comprehensive mutation screening procedure enabled us to identify an NF1 mutation in 93.8% of the NF1 patients in our study. NF1 = neurofibromatosis type 1.

    Article Snippet: To prevent illegitimate splicing, blood samples were processed after venipuncture with a maximum delay of 4 h and samples were not stored at 4°C., Reverse transcription was performed using 500 ng of total RNA isolated and random hexamers with a First-Strand complementary DNA (cDNA) Synthesis Kit for RT-PCR (AMV) (Roche Applied Science, Indianapolis, IN).

    Techniques: Flow Cytometry, Mutagenesis, DNA Sequencing, Sequencing, Multiplex Assay, Ligation, Amplification, Multiplex Ligation-dependent Probe Amplification