dna fragments  (Thermo Fisher)


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    Structured Review

    Thermo Fisher dna fragments
    Semi-quantitative RT-PCR of <t>SLC6A3,</t> BIGH3 and vWF. Total RNA was extracted from 8 pairs of RCC tissue (C) and patient-matched normal kidney tissue (N). The over-expression of SLC6A3 was seen in all 8 tissue pairs and the over-expression of BIGH3 and vWF was seen in 7 of the 8 tissue pairs (sample 1, 3, 4, 5, 6, 7, 8). Amplification of <t>DNA</t> fragment of α-tubulin was used as quantitative control.
    Dna Fragments, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Price from $9.99 to $1999.99
    dna fragments - by Bioz Stars, 2021-03
    86/100 stars

    Images

    1) Product Images from "Microarray gene expression profiling and analysis in renal cell carcinoma"

    Article Title: Microarray gene expression profiling and analysis in renal cell carcinoma

    Journal: BMC Urology

    doi: 10.1186/1471-2490-4-9

    Semi-quantitative RT-PCR of SLC6A3, BIGH3 and vWF. Total RNA was extracted from 8 pairs of RCC tissue (C) and patient-matched normal kidney tissue (N). The over-expression of SLC6A3 was seen in all 8 tissue pairs and the over-expression of BIGH3 and vWF was seen in 7 of the 8 tissue pairs (sample 1, 3, 4, 5, 6, 7, 8). Amplification of DNA fragment of α-tubulin was used as quantitative control.
    Figure Legend Snippet: Semi-quantitative RT-PCR of SLC6A3, BIGH3 and vWF. Total RNA was extracted from 8 pairs of RCC tissue (C) and patient-matched normal kidney tissue (N). The over-expression of SLC6A3 was seen in all 8 tissue pairs and the over-expression of BIGH3 and vWF was seen in 7 of the 8 tissue pairs (sample 1, 3, 4, 5, 6, 7, 8). Amplification of DNA fragment of α-tubulin was used as quantitative control.

    Techniques Used: Quantitative RT-PCR, Over Expression, Amplification

    2) Product Images from "Development of a new marker system for identifying the complex members of the low-molecular-weight glutenin subunit gene family in bread wheat (Triticum aestivum L.)"

    Article Title: Development of a new marker system for identifying the complex members of the low-molecular-weight glutenin subunit gene family in bread wheat (Triticum aestivum L.)

    Journal: TAG. Theoretical and Applied Genetics. Theoretische Und Angewandte Genetik

    doi: 10.1007/s00122-011-1550-7

    Electropherograms displaying the patterns of the DNA fragments detected in Chinese Spring and its nulli–tetrasomic lines with the marker system. a Identification of the LMW-GS genes in Chinese Spring with the developed marker system. b Determination of the location of the DNA fragments with the Chinese Spring nulli–tetrasomic lines N1AT1D, N1BT1D and N1DT1B. The horizontal and vertical axes are the same as those described in Fig. 3 b. In Chinese Spring, 15 blue peaks (DNA fragments) could be detected with the conserved primer LMWGS1, 14 with LMWGS2 and 16 with LMWGS3 (7 with LMWGS3a, 7 with LMWGS3b and 2 with LMWGS3c). The data collected from three sets of conserved primers indicated that 12 blue peaks (DNA fragments) were detected in N1AT1D, 12 in N1BT1D and 9 in N1DT1B. However, DNA fragment 684 (marked with red arrows ) could be detected in all three nulli–tetrasomic lines. All data provided are representative of four independent sets of PCR amplifications and analyses using the Applied Biosystems 3730 DNA Analyzer
    Figure Legend Snippet: Electropherograms displaying the patterns of the DNA fragments detected in Chinese Spring and its nulli–tetrasomic lines with the marker system. a Identification of the LMW-GS genes in Chinese Spring with the developed marker system. b Determination of the location of the DNA fragments with the Chinese Spring nulli–tetrasomic lines N1AT1D, N1BT1D and N1DT1B. The horizontal and vertical axes are the same as those described in Fig. 3 b. In Chinese Spring, 15 blue peaks (DNA fragments) could be detected with the conserved primer LMWGS1, 14 with LMWGS2 and 16 with LMWGS3 (7 with LMWGS3a, 7 with LMWGS3b and 2 with LMWGS3c). The data collected from three sets of conserved primers indicated that 12 blue peaks (DNA fragments) were detected in N1AT1D, 12 in N1BT1D and 9 in N1DT1B. However, DNA fragment 684 (marked with red arrows ) could be detected in all three nulli–tetrasomic lines. All data provided are representative of four independent sets of PCR amplifications and analyses using the Applied Biosystems 3730 DNA Analyzer

    Techniques Used: Marker, Polymerase Chain Reaction

    Analysis of polymerase chain reaction (PCR) products amplified from Xiaoyan 54 with the conserved primers. a Electrophoresis of the PCR products amplified from Xiaoyan 54 with the conserved primers in an agarose gel. M DNA ladder Marker III (200, 500, 800, 1,200, 2,000, 3,000 and 4,500 bp; Tiangen Biotech Co., Ltd.). b Electropherograms showing capillary electrophoresis separation of the DNA fragments amplified from Xiaoyan 54 with the conserved primers. The horizontal axis shows the size of the detected DNA fragments, while the vertical axis displays the intensity of the signal (i.e. the concentration of DNA fragments in the PCR products). The orange peaks match the standard DNA fragments in the GeneScan 1200 LIZ size standard, while the blue ones representing the DNA fragments in the PCR products, should match theoretically with the LMW-GS genes in Xiaoyan 54. The numbers on the horizontal axis represent the size of the corresponding peak in the GeneScan 1200 LIZ size standard ( orange ). In Xiaoyan 54, 14 blue peaks (DNA fragments) could be detected with the conserved primer LMWGS1, 16 with LMWGS2 and 16 with LMWGS3 (7 with LMWGS3a, 6 with LMWGS3b and 3 with LMWGS3c). The data shown are representative of four independent sets of PCR amplifications and DNA fragment analyses
    Figure Legend Snippet: Analysis of polymerase chain reaction (PCR) products amplified from Xiaoyan 54 with the conserved primers. a Electrophoresis of the PCR products amplified from Xiaoyan 54 with the conserved primers in an agarose gel. M DNA ladder Marker III (200, 500, 800, 1,200, 2,000, 3,000 and 4,500 bp; Tiangen Biotech Co., Ltd.). b Electropherograms showing capillary electrophoresis separation of the DNA fragments amplified from Xiaoyan 54 with the conserved primers. The horizontal axis shows the size of the detected DNA fragments, while the vertical axis displays the intensity of the signal (i.e. the concentration of DNA fragments in the PCR products). The orange peaks match the standard DNA fragments in the GeneScan 1200 LIZ size standard, while the blue ones representing the DNA fragments in the PCR products, should match theoretically with the LMW-GS genes in Xiaoyan 54. The numbers on the horizontal axis represent the size of the corresponding peak in the GeneScan 1200 LIZ size standard ( orange ). In Xiaoyan 54, 14 blue peaks (DNA fragments) could be detected with the conserved primer LMWGS1, 16 with LMWGS2 and 16 with LMWGS3 (7 with LMWGS3a, 6 with LMWGS3b and 3 with LMWGS3c). The data shown are representative of four independent sets of PCR amplifications and DNA fragment analyses

    Techniques Used: Polymerase Chain Reaction, Amplification, Electrophoresis, Agarose Gel Electrophoresis, Marker, Concentration Assay

    Related Articles

    Sonication:

    Article Title: Nematode Genome Announcement: A Draft Genome for Rice Root-Knot Nematode, Meloidogyne graminicola
    Article Snippet: Freshly hatched second stage juveniles were used for the genomic DNA extraction using Gentra Puregene Tissue Kit (Cat No.: 158667 Qiagen, Valencia, CA, USA). .. The short (150–200 bp) and long (300–500 bp) DNA fragments were obtained by diluting 1 µg of genomic DNA in 100 µl nuclease free water (Ambion, Waltham, MA, USA) and sonication by Bioruptor (Diagenode, Seraing (Ougrée), Belgium) at 20 and 13 pulses at 30 sec ON and 30 sec OFF, respectively. .. The resulting fragmented DNA was cleaned using QIAquick columns (Qiagen, Valencia, CA, USA).

    Article Title: Epigenetic regulation of olfactory receptor gene expression by the Myb-MuvB/dREAM complex
    Article Snippet: .. Chromatin was sonicated to obtain 300- to 500-base-pair (bp) DNA fragments and incubated with Dynabeads (Invitrogen) and 2 μg of antibodies to H3K9me2 (ab1220) or H3K4me3 (ab8580). .. Specificity of anti-H3K9me2 antibody was validated using a peptide array (Supplemental Fig. S7A).

    Polymerase Chain Reaction:

    Article Title: SCYL2 Genes Are Involved in Clathrin-Mediated Vesicle Trafficking and Essential for Plant Growth 1SCYL2 Genes Are Involved in Clathrin-Mediated Vesicle Trafficking and Essential for Plant Growth 1 [OPEN]
    Article Snippet: To generate the pMDC43 : EXPA7promoter : N-GFP plasmids carrying various N-terminal GFP-fused genes that encode proteins with root hair tip localization, the 35S promoter of pMDC43 vector was replaced with the 1.5-kb promoter of EXPA7 by the recombination of pMDC43 DNA fragments digested with Hin dIII and Kpn I and PCR-amplified DNA fragments using primers 5′-CGACGGCCAGTGCCAAAGCTTAATTAGGGTCCAAGGTTTGTTC-3′ and 5′-CATTTTTTCTACCGGTACCTCTAGCCTCTTTTTCTTTATTCTTA-3′. .. Then, the open reading frames of full-length genes of ROP2, RABA4B, RHD2, CSLD3, SYP123, and PRP3 were PCR amplified using the following primers: sense 5′-CCGGTACCGAATTCGATGGCGTCAAGGTTTATAAAGTGT-3′ and antisense 5′-ATATCTCGAGTGCGGTCACAAGAACGCGCAACGGTTCTT-3′ for ROP2 (AT1G20090), sense 5′-CCGGTACCGAATTCGATGGCCGGAGGAGGCGGATAC-3′ and antisense 5′-ATATCTCGAGTGCGGTCAAGAAGAAGTACAACAAGTGCT-3′ for RABA4B (AT4G39990), sense 5′-CCGGTACCGAATTCGATGTCTAGAGTGAGTTTTGAAGTG-3′ and antisense 5′-ATATCTCGAGTGCGGTTAGAAATTCTCTTTGTGGAAGGA-3′ for RHD2 (AT5G51060), sense 5′-CCGGTACCGAATTCGATGGCGTCTAATAATCATTTCATG-3′ and antisense 5′-ATATCTCGAGTGCGGTCATGGGAAAGTGAAAGATCCTCC-3′ for CSLD3 (AT3G03050), sense 5′-CCGGTACCGAATTCGATGAACGATCTTATCTCAAGCTCA-3′ and antisense 5′-ATATCTCGAGTGCGGTCAAGGTCGAAGTAGAGTGTTAAA-3′ for SYP123 (AT4G03330), and sense 5′-CCGGTACCGAATTCGATGGCGATCACACGCTCCTCCT-3′ and antisense 5′-ATATCTCGAGTGCGGTCAGTATTTGGGAGTGGCGGG-3′ for PRP3 (AT3G62680), and recombined with pENTR2B (Invitrogen) DNA fragments amplified by PCR using primers 5′-CCGCACTCGAGATATCTAGAC-3′ and 5′-CGAATTCGGTACCGGATC-3′. .. The resulting pENTR2B vectors with genes were recombined with the destination plasmid pMDC43 : EXPA7promoter to yield the GFP fusion genes in pMDC43 : EXPA7promoter plasmids, and T2 transgenic plants in the background of and scyl2b mutants were generated and used for the root hair tip localization study.

    Article Title: Distinct Stage Changes in Early-Life Colonization and Acquisition of the Gut Microbiota and Its Correlations With Volatile Fatty Acids in Goat Kids
    Article Snippet: High-Throughput Sequencing and Data Analysis To analyze the bacterial communities, the V3–V4 region of the 16S rRNA gene was amplified using a primer pair (341F: 5′-CCTAYGGGRBGCASCAG-3′ and 806R: 5′-GGA CTACNNGGGTATCTAAT-3′) , and the thermocycling protocol of the amplification was: 98°C for 1 min, followed by 30 cycles at 98°C for 10 s, 50°C for 30 s, and 72°C for 30 s, followed by a final extension at 72°C for 5 min. .. The PCR products (DNA fragments of 450–550 bp in length) were purified by 2% agarose gel electrophoresis and the GeneJET Gel Extraction Kit (Thermo Fisher Scientific, United States) before the construction of sequencing libraries. .. All libraries (n = 123) were subjected to 2 × 250 paired-end sequencing on an Illumina NovaSeq 6000 platform (Novogene Co., Ltd., Beijing, China).

    Article Title: Both microsatellite length and sequence context determine frameshift mutation rates in defective DNA mismatch repair
    Article Snippet: The PCR products were used for DNA sequencing to identify stable cell lines and frameshift mutations at microsatellites of TGFBR2 and ACVR2 . .. In addition, we subcloned PCR-amplified TGFBR2 OF (A13 ) and ACVR2 OF (A13 ) DNA fragments from the M1 and M2 cell populations by utilizing a TA cloning vector (Invitrogen Corp.) as per the manufacturer's protocol. .. DNA clones were then individually sequenced to determine the prevalence of mutated and A13 microsatellite sequences of TGFBR2 and ACVR2 .

    Article Title: Analysis of the leakage of gene repression by an artificial TetR-regulated promoter in cyanobacteria
    Article Snippet: .. DNA fragments DNA fragments comprising a L promoter were prepared in length of 160 bp (Fig. ) and biotinylated at the 5′-end of the template strand by PCR using the Phusion Hot Start II High-Fidelity DNA Polymerase (Thermo Scientific, Waltham, USA) with the forward and the biotinylated reverse primers. ..

    Amplification:

    Article Title: SCYL2 Genes Are Involved in Clathrin-Mediated Vesicle Trafficking and Essential for Plant Growth 1SCYL2 Genes Are Involved in Clathrin-Mediated Vesicle Trafficking and Essential for Plant Growth 1 [OPEN]
    Article Snippet: To generate the pMDC43 : EXPA7promoter : N-GFP plasmids carrying various N-terminal GFP-fused genes that encode proteins with root hair tip localization, the 35S promoter of pMDC43 vector was replaced with the 1.5-kb promoter of EXPA7 by the recombination of pMDC43 DNA fragments digested with Hin dIII and Kpn I and PCR-amplified DNA fragments using primers 5′-CGACGGCCAGTGCCAAAGCTTAATTAGGGTCCAAGGTTTGTTC-3′ and 5′-CATTTTTTCTACCGGTACCTCTAGCCTCTTTTTCTTTATTCTTA-3′. .. Then, the open reading frames of full-length genes of ROP2, RABA4B, RHD2, CSLD3, SYP123, and PRP3 were PCR amplified using the following primers: sense 5′-CCGGTACCGAATTCGATGGCGTCAAGGTTTATAAAGTGT-3′ and antisense 5′-ATATCTCGAGTGCGGTCACAAGAACGCGCAACGGTTCTT-3′ for ROP2 (AT1G20090), sense 5′-CCGGTACCGAATTCGATGGCCGGAGGAGGCGGATAC-3′ and antisense 5′-ATATCTCGAGTGCGGTCAAGAAGAAGTACAACAAGTGCT-3′ for RABA4B (AT4G39990), sense 5′-CCGGTACCGAATTCGATGTCTAGAGTGAGTTTTGAAGTG-3′ and antisense 5′-ATATCTCGAGTGCGGTTAGAAATTCTCTTTGTGGAAGGA-3′ for RHD2 (AT5G51060), sense 5′-CCGGTACCGAATTCGATGGCGTCTAATAATCATTTCATG-3′ and antisense 5′-ATATCTCGAGTGCGGTCATGGGAAAGTGAAAGATCCTCC-3′ for CSLD3 (AT3G03050), sense 5′-CCGGTACCGAATTCGATGAACGATCTTATCTCAAGCTCA-3′ and antisense 5′-ATATCTCGAGTGCGGTCAAGGTCGAAGTAGAGTGTTAAA-3′ for SYP123 (AT4G03330), and sense 5′-CCGGTACCGAATTCGATGGCGATCACACGCTCCTCCT-3′ and antisense 5′-ATATCTCGAGTGCGGTCAGTATTTGGGAGTGGCGGG-3′ for PRP3 (AT3G62680), and recombined with pENTR2B (Invitrogen) DNA fragments amplified by PCR using primers 5′-CCGCACTCGAGATATCTAGAC-3′ and 5′-CGAATTCGGTACCGGATC-3′. .. The resulting pENTR2B vectors with genes were recombined with the destination plasmid pMDC43 : EXPA7promoter to yield the GFP fusion genes in pMDC43 : EXPA7promoter plasmids, and T2 transgenic plants in the background of and scyl2b mutants were generated and used for the root hair tip localization study.

    Article Title: Deleterious genetic variants in ciliopathy genes increase risk of ritodrine-induced cardiac and pulmonary side effects
    Article Snippet: .. WES and variant calls Genomic DNA extracted from peripheral blood cells was amplified to generate 175–250-base pair (bp) DNA fragments spanning the protein-coding regions of human genome DNA using the Ion AmpliSeq Exome Panel (Thermo Fisher Scientific, Waltham, MA, USA). .. Library construction was performed to load the DNA samples into the semiconductor chip using the Ion AmpliSeq Exome Library Kit Plus, covering 57,742,646 bp (1.85% of human genomic regions) as described in the manufacturer’s instructions (Thermo Fisher Scientific).

    Article Title: Deleterious genetic variants in ciliopathy genes increase risk of ritodrine-induced cardiac and pulmonary side effects
    Article Snippet: .. Genomic DNA extracted from peripheral blood cells was amplified to generate 175–250-base pair (bp) DNA fragments spanning the protein-coding regions of human genome DNA using the Ion AmpliSeq Exome Panel (Thermo Fisher Scientific, Waltham, MA, USA). .. Library construction was performed to load the DNA samples into the semiconductor chip using the Ion AmpliSeq Exome Library Kit Plus, covering 57,742,646 bp (1.85% of human genomic regions) as described in the manufacturer’s instructions (Thermo Fisher Scientific).

    Variant Assay:

    Article Title: Deleterious genetic variants in ciliopathy genes increase risk of ritodrine-induced cardiac and pulmonary side effects
    Article Snippet: .. WES and variant calls Genomic DNA extracted from peripheral blood cells was amplified to generate 175–250-base pair (bp) DNA fragments spanning the protein-coding regions of human genome DNA using the Ion AmpliSeq Exome Panel (Thermo Fisher Scientific, Waltham, MA, USA). .. Library construction was performed to load the DNA samples into the semiconductor chip using the Ion AmpliSeq Exome Library Kit Plus, covering 57,742,646 bp (1.85% of human genomic regions) as described in the manufacturer’s instructions (Thermo Fisher Scientific).

    Incubation:

    Article Title: Epigenetic regulation of olfactory receptor gene expression by the Myb-MuvB/dREAM complex
    Article Snippet: .. Chromatin was sonicated to obtain 300- to 500-base-pair (bp) DNA fragments and incubated with Dynabeads (Invitrogen) and 2 μg of antibodies to H3K9me2 (ab1220) or H3K4me3 (ab8580). .. Specificity of anti-H3K9me2 antibody was validated using a peptide array (Supplemental Fig. S7A).

    Purification:

    Article Title: Distinct Stage Changes in Early-Life Colonization and Acquisition of the Gut Microbiota and Its Correlations With Volatile Fatty Acids in Goat Kids
    Article Snippet: High-Throughput Sequencing and Data Analysis To analyze the bacterial communities, the V3–V4 region of the 16S rRNA gene was amplified using a primer pair (341F: 5′-CCTAYGGGRBGCASCAG-3′ and 806R: 5′-GGA CTACNNGGGTATCTAAT-3′) , and the thermocycling protocol of the amplification was: 98°C for 1 min, followed by 30 cycles at 98°C for 10 s, 50°C for 30 s, and 72°C for 30 s, followed by a final extension at 72°C for 5 min. .. The PCR products (DNA fragments of 450–550 bp in length) were purified by 2% agarose gel electrophoresis and the GeneJET Gel Extraction Kit (Thermo Fisher Scientific, United States) before the construction of sequencing libraries. .. All libraries (n = 123) were subjected to 2 × 250 paired-end sequencing on an Illumina NovaSeq 6000 platform (Novogene Co., Ltd., Beijing, China).

    Agarose Gel Electrophoresis:

    Article Title: Distinct Stage Changes in Early-Life Colonization and Acquisition of the Gut Microbiota and Its Correlations With Volatile Fatty Acids in Goat Kids
    Article Snippet: High-Throughput Sequencing and Data Analysis To analyze the bacterial communities, the V3–V4 region of the 16S rRNA gene was amplified using a primer pair (341F: 5′-CCTAYGGGRBGCASCAG-3′ and 806R: 5′-GGA CTACNNGGGTATCTAAT-3′) , and the thermocycling protocol of the amplification was: 98°C for 1 min, followed by 30 cycles at 98°C for 10 s, 50°C for 30 s, and 72°C for 30 s, followed by a final extension at 72°C for 5 min. .. The PCR products (DNA fragments of 450–550 bp in length) were purified by 2% agarose gel electrophoresis and the GeneJET Gel Extraction Kit (Thermo Fisher Scientific, United States) before the construction of sequencing libraries. .. All libraries (n = 123) were subjected to 2 × 250 paired-end sequencing on an Illumina NovaSeq 6000 platform (Novogene Co., Ltd., Beijing, China).

    Gel Extraction:

    Article Title: Distinct Stage Changes in Early-Life Colonization and Acquisition of the Gut Microbiota and Its Correlations With Volatile Fatty Acids in Goat Kids
    Article Snippet: High-Throughput Sequencing and Data Analysis To analyze the bacterial communities, the V3–V4 region of the 16S rRNA gene was amplified using a primer pair (341F: 5′-CCTAYGGGRBGCASCAG-3′ and 806R: 5′-GGA CTACNNGGGTATCTAAT-3′) , and the thermocycling protocol of the amplification was: 98°C for 1 min, followed by 30 cycles at 98°C for 10 s, 50°C for 30 s, and 72°C for 30 s, followed by a final extension at 72°C for 5 min. .. The PCR products (DNA fragments of 450–550 bp in length) were purified by 2% agarose gel electrophoresis and the GeneJET Gel Extraction Kit (Thermo Fisher Scientific, United States) before the construction of sequencing libraries. .. All libraries (n = 123) were subjected to 2 × 250 paired-end sequencing on an Illumina NovaSeq 6000 platform (Novogene Co., Ltd., Beijing, China).

    Sequencing:

    Article Title: Distinct Stage Changes in Early-Life Colonization and Acquisition of the Gut Microbiota and Its Correlations With Volatile Fatty Acids in Goat Kids
    Article Snippet: High-Throughput Sequencing and Data Analysis To analyze the bacterial communities, the V3–V4 region of the 16S rRNA gene was amplified using a primer pair (341F: 5′-CCTAYGGGRBGCASCAG-3′ and 806R: 5′-GGA CTACNNGGGTATCTAAT-3′) , and the thermocycling protocol of the amplification was: 98°C for 1 min, followed by 30 cycles at 98°C for 10 s, 50°C for 30 s, and 72°C for 30 s, followed by a final extension at 72°C for 5 min. .. The PCR products (DNA fragments of 450–550 bp in length) were purified by 2% agarose gel electrophoresis and the GeneJET Gel Extraction Kit (Thermo Fisher Scientific, United States) before the construction of sequencing libraries. .. All libraries (n = 123) were subjected to 2 × 250 paired-end sequencing on an Illumina NovaSeq 6000 platform (Novogene Co., Ltd., Beijing, China).

    TA Cloning:

    Article Title: Both microsatellite length and sequence context determine frameshift mutation rates in defective DNA mismatch repair
    Article Snippet: The PCR products were used for DNA sequencing to identify stable cell lines and frameshift mutations at microsatellites of TGFBR2 and ACVR2 . .. In addition, we subcloned PCR-amplified TGFBR2 OF (A13 ) and ACVR2 OF (A13 ) DNA fragments from the M1 and M2 cell populations by utilizing a TA cloning vector (Invitrogen Corp.) as per the manufacturer's protocol. .. DNA clones were then individually sequenced to determine the prevalence of mutated and A13 microsatellite sequences of TGFBR2 and ACVR2 .

    Plasmid Preparation:

    Article Title: Both microsatellite length and sequence context determine frameshift mutation rates in defective DNA mismatch repair
    Article Snippet: The PCR products were used for DNA sequencing to identify stable cell lines and frameshift mutations at microsatellites of TGFBR2 and ACVR2 . .. In addition, we subcloned PCR-amplified TGFBR2 OF (A13 ) and ACVR2 OF (A13 ) DNA fragments from the M1 and M2 cell populations by utilizing a TA cloning vector (Invitrogen Corp.) as per the manufacturer's protocol. .. DNA clones were then individually sequenced to determine the prevalence of mutated and A13 microsatellite sequences of TGFBR2 and ACVR2 .

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    Thermo Fisher biotinylated dna fragments
    ME-Scan library preparation ( A–E ) and sequencing ( F–H ). ( A ) Genomic <t>DNA</t> (green) containing an Alu Yb8/9 (blue) in the forward orientation. The 5′ GC-rich region, the 3′ poly-A tail, and the Alu Yb8/9-specific primer site ( Alu BP2) are shown as darker segments. The target site duplications (TSD) are indicated by boxed black arrows. ( B ) DNA samples are fragmented, end-repaired, and an overhanging 3′ A is added. ( C ) Partially double-stranded oligonucleotide adapters (orange) with 9-bp indexes (darker segment) are ligated onto fragment ends. Indexed samples are then pooled. ( D ) Alu Yb8/9 element junctions are targeted by PCR using a <t>biotinylated</t> Alu Yb8/9-specific primer ( Alu BP2) and adapter primer PEP2. Biotinylated DNA molecules are then purified using streptavidin-coated paramagnetic beads. ( E ) Reamplification of the library with primers PEP1 and PEP2 (orange arrows). ( F–H ) The pooled junction library is then sequenced with a three-read design. Primers are shown as arrows, sequencing reads as dashed lines. The first sequencing read (50 nt) extends from Alu SPv2 ( F ) or Alu SPv3 ( G ). To skip the 5′ end of the Alu insertions, which would be identical over the entire flow cell and therefore difficult for Illumina's software to process, the first 30 cycles of nucleotide synthesis are carried out without collecting data (represented by a dashed gray ‘hop’). The second read proceeds for 57 bp from Alu SPIn1. The third read is generated using Illumina's standard second-end read primer and consists of a 9-bp index, a ‘T,’ and 36 bp of genomic sequence from a fragmentation site 50–300 bp upstream of the Alu insertion. Read sets are generated from multiple different fragments representing each Alu Yb8/9 insertion in the library ( H ). Each insertion is uniquely identified by its “ Alu Junction Position” (dashed line and large arrow in F and G ).
    Biotinylated Dna Fragments, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Thermo Fisher mutational analysis dna
    p53 immunohistochemistry of the two remaining discordant cases after consensus. NA, not applicable. For the first case (upper panel), consensus was reached on a mid‐epithelial p53 immunohistochemistry (IHC) expression pattern with basal sparing (indicated by arrow), while the tumour showed a pathogenic TP53 mutation with a high variant allele frequency (VAF). An additional p16–IHC could prevent misinterpretation of the final p53 IHC class due to the absence of ‘block‐type’ p16 expression. Because of suboptimal staining, an alternative block of the same case was stained for p53 and a diffuse expression of p53 was observed. Moreover, after revising all haematoxylin and eosin (H E) staining of this tumour revealed the presence of differentiated type, vulvar intra‐epithelial neoplasia (dVIN) as a precursor lesion, which increases the probability of harbouring TP53 mutations. The second case (lower panel) was also human papillomavirus (HPV)‐unrelated, and scored scattered by both observers. We cannot explain the discordancy between the final p53 IHC class and <t>mutational</t> <t>analysis</t> of this case, although the VAF was low but reliable. Scale bar 20 µm.
    Mutational Analysis Dna, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher genomic dna fragments
    Experimental protocol design. a Scheme of <t>SVA</t> element structure. b Sequence alignment of SVA_1 and SVA_2 primer binding sites and SVA, Alu subfamily consensuses. The SVA_1 and SVA_2 primer sequences are shown above of the alignment and the amplification directions are indicated by arrows. Top row of the sequence alignment shows the sequences of the primer binding sites of SVA_1 and SVA_2. SVA_1 binding site includes the SVA characteristic deletion as compared to Alu sequences. Dots in the alignment represent the same nucleotides as the primer binding site sequences. Deletions are shown as dashes and mutations are shown as the correct base for the consensus. c SVA-specific amplifications during ME-Scan-SVA library construction and the final <t>DNA</t> fragment structure. The DNA library after second-round amplification is size-selected at ~500 bp (an example electropherogram image is shown). White box: adaptor; grey box: index; dark green box: flanking genomic region; yellow box: TSD; orange box: (CCCTCT) n hexamer simple repeat; light green box: SVA Alu -like region
    Genomic Dna Fragments, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ME-Scan library preparation ( A–E ) and sequencing ( F–H ). ( A ) Genomic DNA (green) containing an Alu Yb8/9 (blue) in the forward orientation. The 5′ GC-rich region, the 3′ poly-A tail, and the Alu Yb8/9-specific primer site ( Alu BP2) are shown as darker segments. The target site duplications (TSD) are indicated by boxed black arrows. ( B ) DNA samples are fragmented, end-repaired, and an overhanging 3′ A is added. ( C ) Partially double-stranded oligonucleotide adapters (orange) with 9-bp indexes (darker segment) are ligated onto fragment ends. Indexed samples are then pooled. ( D ) Alu Yb8/9 element junctions are targeted by PCR using a biotinylated Alu Yb8/9-specific primer ( Alu BP2) and adapter primer PEP2. Biotinylated DNA molecules are then purified using streptavidin-coated paramagnetic beads. ( E ) Reamplification of the library with primers PEP1 and PEP2 (orange arrows). ( F–H ) The pooled junction library is then sequenced with a three-read design. Primers are shown as arrows, sequencing reads as dashed lines. The first sequencing read (50 nt) extends from Alu SPv2 ( F ) or Alu SPv3 ( G ). To skip the 5′ end of the Alu insertions, which would be identical over the entire flow cell and therefore difficult for Illumina's software to process, the first 30 cycles of nucleotide synthesis are carried out without collecting data (represented by a dashed gray ‘hop’). The second read proceeds for 57 bp from Alu SPIn1. The third read is generated using Illumina's standard second-end read primer and consists of a 9-bp index, a ‘T,’ and 36 bp of genomic sequence from a fragmentation site 50–300 bp upstream of the Alu insertion. Read sets are generated from multiple different fragments representing each Alu Yb8/9 insertion in the library ( H ). Each insertion is uniquely identified by its “ Alu Junction Position” (dashed line and large arrow in F and G ).

    Journal: Genome Research

    Article Title: Mobile element scanning (ME-Scan) identifies thousands of novel Alu insertions in diverse human populations

    doi: 10.1101/gr.148973.112

    Figure Lengend Snippet: ME-Scan library preparation ( A–E ) and sequencing ( F–H ). ( A ) Genomic DNA (green) containing an Alu Yb8/9 (blue) in the forward orientation. The 5′ GC-rich region, the 3′ poly-A tail, and the Alu Yb8/9-specific primer site ( Alu BP2) are shown as darker segments. The target site duplications (TSD) are indicated by boxed black arrows. ( B ) DNA samples are fragmented, end-repaired, and an overhanging 3′ A is added. ( C ) Partially double-stranded oligonucleotide adapters (orange) with 9-bp indexes (darker segment) are ligated onto fragment ends. Indexed samples are then pooled. ( D ) Alu Yb8/9 element junctions are targeted by PCR using a biotinylated Alu Yb8/9-specific primer ( Alu BP2) and adapter primer PEP2. Biotinylated DNA molecules are then purified using streptavidin-coated paramagnetic beads. ( E ) Reamplification of the library with primers PEP1 and PEP2 (orange arrows). ( F–H ) The pooled junction library is then sequenced with a three-read design. Primers are shown as arrows, sequencing reads as dashed lines. The first sequencing read (50 nt) extends from Alu SPv2 ( F ) or Alu SPv3 ( G ). To skip the 5′ end of the Alu insertions, which would be identical over the entire flow cell and therefore difficult for Illumina's software to process, the first 30 cycles of nucleotide synthesis are carried out without collecting data (represented by a dashed gray ‘hop’). The second read proceeds for 57 bp from Alu SPIn1. The third read is generated using Illumina's standard second-end read primer and consists of a 9-bp index, a ‘T,’ and 36 bp of genomic sequence from a fragmentation site 50–300 bp upstream of the Alu insertion. Read sets are generated from multiple different fragments representing each Alu Yb8/9 insertion in the library ( H ). Each insertion is uniquely identified by its “ Alu Junction Position” (dashed line and large arrow in F and G ).

    Article Snippet: The size-selected DNA was incubated with streptavidin-coated paramagnetic beads to retain biotinylated DNA fragments per the manufacturer's protocol (Dynabeads MyOne Streptavidin C1, Life Technologies, Inc.).

    Techniques: Sequencing, Polymerase Chain Reaction, Purification, Flow Cytometry, Software, Generated

    p53 immunohistochemistry of the two remaining discordant cases after consensus. NA, not applicable. For the first case (upper panel), consensus was reached on a mid‐epithelial p53 immunohistochemistry (IHC) expression pattern with basal sparing (indicated by arrow), while the tumour showed a pathogenic TP53 mutation with a high variant allele frequency (VAF). An additional p16–IHC could prevent misinterpretation of the final p53 IHC class due to the absence of ‘block‐type’ p16 expression. Because of suboptimal staining, an alternative block of the same case was stained for p53 and a diffuse expression of p53 was observed. Moreover, after revising all haematoxylin and eosin (H E) staining of this tumour revealed the presence of differentiated type, vulvar intra‐epithelial neoplasia (dVIN) as a precursor lesion, which increases the probability of harbouring TP53 mutations. The second case (lower panel) was also human papillomavirus (HPV)‐unrelated, and scored scattered by both observers. We cannot explain the discordancy between the final p53 IHC class and mutational analysis of this case, although the VAF was low but reliable. Scale bar 20 µm.

    Journal: Histopathology

    Article Title: Performance of the pattern‐based interpretation of p53 immunohistochemistry as a surrogate for TP53 mutations in vulvar squamous cell carcinomaPerformance of the pattern‐based interpretation of p53 immunohistochemistry as a surrogate for TP53 mutations in vulvar squamous cell carcinoma

    doi: 10.1111/his.14109

    Figure Lengend Snippet: p53 immunohistochemistry of the two remaining discordant cases after consensus. NA, not applicable. For the first case (upper panel), consensus was reached on a mid‐epithelial p53 immunohistochemistry (IHC) expression pattern with basal sparing (indicated by arrow), while the tumour showed a pathogenic TP53 mutation with a high variant allele frequency (VAF). An additional p16–IHC could prevent misinterpretation of the final p53 IHC class due to the absence of ‘block‐type’ p16 expression. Because of suboptimal staining, an alternative block of the same case was stained for p53 and a diffuse expression of p53 was observed. Moreover, after revising all haematoxylin and eosin (H E) staining of this tumour revealed the presence of differentiated type, vulvar intra‐epithelial neoplasia (dVIN) as a precursor lesion, which increases the probability of harbouring TP53 mutations. The second case (lower panel) was also human papillomavirus (HPV)‐unrelated, and scored scattered by both observers. We cannot explain the discordancy between the final p53 IHC class and mutational analysis of this case, although the VAF was low but reliable. Scale bar 20 µm.

    Article Snippet: MUTATIONAL ANALYSIS DNA was isolated locally from each case using different techniques.

    Techniques: Immunohistochemistry, Expressing, Mutagenesis, Variant Assay, Blocking Assay, Staining

    Experimental protocol design. a Scheme of SVA element structure. b Sequence alignment of SVA_1 and SVA_2 primer binding sites and SVA, Alu subfamily consensuses. The SVA_1 and SVA_2 primer sequences are shown above of the alignment and the amplification directions are indicated by arrows. Top row of the sequence alignment shows the sequences of the primer binding sites of SVA_1 and SVA_2. SVA_1 binding site includes the SVA characteristic deletion as compared to Alu sequences. Dots in the alignment represent the same nucleotides as the primer binding site sequences. Deletions are shown as dashes and mutations are shown as the correct base for the consensus. c SVA-specific amplifications during ME-Scan-SVA library construction and the final DNA fragment structure. The DNA library after second-round amplification is size-selected at ~500 bp (an example electropherogram image is shown). White box: adaptor; grey box: index; dark green box: flanking genomic region; yellow box: TSD; orange box: (CCCTCT) n hexamer simple repeat; light green box: SVA Alu -like region

    Journal: Mobile DNA

    Article Title: Identification of polymorphic SVA retrotransposons using a mobile element scanning method for SVA (ME-Scan-SVA)

    doi: 10.1186/s13100-016-0072-x

    Figure Lengend Snippet: Experimental protocol design. a Scheme of SVA element structure. b Sequence alignment of SVA_1 and SVA_2 primer binding sites and SVA, Alu subfamily consensuses. The SVA_1 and SVA_2 primer sequences are shown above of the alignment and the amplification directions are indicated by arrows. Top row of the sequence alignment shows the sequences of the primer binding sites of SVA_1 and SVA_2. SVA_1 binding site includes the SVA characteristic deletion as compared to Alu sequences. Dots in the alignment represent the same nucleotides as the primer binding site sequences. Deletions are shown as dashes and mutations are shown as the correct base for the consensus. c SVA-specific amplifications during ME-Scan-SVA library construction and the final DNA fragment structure. The DNA library after second-round amplification is size-selected at ~500 bp (an example electropherogram image is shown). White box: adaptor; grey box: index; dark green box: flanking genomic region; yellow box: TSD; orange box: (CCCTCT) n hexamer simple repeat; light green box: SVA Alu -like region

    Article Snippet: After size selection, biotinylated SVA-enriched DNA fragments were magnetically separated from other genomic DNA fragments using 5 μl DynabeadsR M-270 Streptavidin (cat. no. 65305, Invitrogen, Life Technologies, Oslo, Norway) following the manufacturer’s protocol.

    Techniques: Sequencing, Binding Assay, Amplification