rna  (Qiagen)


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    Name:
    AllPrep DNA RNA Mini Kit
    Description:
    For simultaneous purification of DNA and RNA from cells and tissues Kit contents Qiagen AllPrep DNA RNA Mini Kit 50 preps 30mg Sample 100L Elution Volume Silica Technology Spin Column Format Manual Processing Genomic DNA Total RNA Purification 35 min Time Run Ideal for PCR Real time PCR Microarray Blotting For Simultaneous Purification of DNA and RNA from Cells and Tissues Includes AllPrep DNA Spin Columns RNeasy Mini Spin Columns Collection Tubes RNase free Water and Buffers Benefits High quality DNA and RNA from the same sample Maximal yields of DNA and RNA from precious samples Rapid purification with short streamlined protocol Ready to use DNA and RNA for any downstream analysis
    Catalog Number:
    80204
    Price:
    541
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    AllPrep DNA RNA Mini Kit
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    Structured Review

    Qiagen rna
    AllPrep DNA RNA Mini Kit
    For simultaneous purification of DNA and RNA from cells and tissues Kit contents Qiagen AllPrep DNA RNA Mini Kit 50 preps 30mg Sample 100L Elution Volume Silica Technology Spin Column Format Manual Processing Genomic DNA Total RNA Purification 35 min Time Run Ideal for PCR Real time PCR Microarray Blotting For Simultaneous Purification of DNA and RNA from Cells and Tissues Includes AllPrep DNA Spin Columns RNeasy Mini Spin Columns Collection Tubes RNase free Water and Buffers Benefits High quality DNA and RNA from the same sample Maximal yields of DNA and RNA from precious samples Rapid purification with short streamlined protocol Ready to use DNA and RNA for any downstream analysis
    https://www.bioz.com/result/rna/product/Qiagen
    Average 97 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rna - by Bioz Stars, 2021-03
    97/100 stars

    Images

    1) Product Images from "Chromatin maturation of the HIV-1 provirus in primary resting CD4+ T cells"

    Article Title: Chromatin maturation of the HIV-1 provirus in primary resting CD4+ T cells

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1008264

    Primary cells to study long-term HIV-1 latency establishment. ( A ) Schematic of the generation of HIV-1 carrying Bcl2 model cells. ( B ). ddPCR scatterplots of HIV-1 infected Bcl2-cells from three HIV-negative donors with probes against env (y-axis) and gag (x-axis). DNA isolated at 3 dpi. ( C ) Quantification of ddPCR results over three probes ( 5´LTR , gag and env ) in control cell line J-lat 5A8, and primary cells from the three donors in biological duplicate, either with both Bcl2 and HIV-1 or Bcl2-only. The data from individual donors and duplicates are visualized by differently shaped points. ( D ) Ratios of gag / env ddPCR signals to estimate internal 5´and 3´ HIV-1 deletions in the three donors and the control cell line J-lat 5A8. ( E ) Fraction of “rain”, i.e. low env signals (approximately below 10,000 a.u.) reflecting APOBEC3G-induced hypermutations. ( F ) RT-ddPCR results of cell-associated RNA isolated at 3 dpi ( n = 2). Probes were as in previous results. *p
    Figure Legend Snippet: Primary cells to study long-term HIV-1 latency establishment. ( A ) Schematic of the generation of HIV-1 carrying Bcl2 model cells. ( B ). ddPCR scatterplots of HIV-1 infected Bcl2-cells from three HIV-negative donors with probes against env (y-axis) and gag (x-axis). DNA isolated at 3 dpi. ( C ) Quantification of ddPCR results over three probes ( 5´LTR , gag and env ) in control cell line J-lat 5A8, and primary cells from the three donors in biological duplicate, either with both Bcl2 and HIV-1 or Bcl2-only. The data from individual donors and duplicates are visualized by differently shaped points. ( D ) Ratios of gag / env ddPCR signals to estimate internal 5´and 3´ HIV-1 deletions in the three donors and the control cell line J-lat 5A8. ( E ) Fraction of “rain”, i.e. low env signals (approximately below 10,000 a.u.) reflecting APOBEC3G-induced hypermutations. ( F ) RT-ddPCR results of cell-associated RNA isolated at 3 dpi ( n = 2). Probes were as in previous results. *p

    Techniques Used: Infection, Isolation

    2) Product Images from "Minocycline Attenuates HIV Infection and Reactivation by Suppressing Cellular Activation in Human CD4+ T Cells"

    Article Title: Minocycline Attenuates HIV Infection and Reactivation by Suppressing Cellular Activation in Human CD4+ T Cells

    Journal: The Journal of Infectious Diseases

    doi: 10.1086/651277

    Intracellular human immunodeficiency virus (HIV) DNA and RNA levels after in vitro HIV NL4-3 infection of CD4 + T cells. CD4 + T cells were pretreated with minocycline ( diamonds ) or not pretreated ( squares ), followed by activation with anti-CD3/CD28 antibodies
    Figure Legend Snippet: Intracellular human immunodeficiency virus (HIV) DNA and RNA levels after in vitro HIV NL4-3 infection of CD4 + T cells. CD4 + T cells were pretreated with minocycline ( diamonds ) or not pretreated ( squares ), followed by activation with anti-CD3/CD28 antibodies

    Techniques Used: In Vitro, Infection, Activation Assay

    Related Articles

    Lysis:

    Article Title: Seasonal dynamics of active SAR11 ecotypes in the oligotrophic Northwest Mediterranean Sea
    Article Snippet: .. A combination of mechanic (freeze-thaw) and enzymatic cell lysis techniques were applied directly to Sterivex cartridges, followed by extraction using an AllPrep DNA/RNA kit (Qiagen, Venlo, Netherlands). .. The RNA samples were tested for the presence of contaminating genomic DNA by PCR and then reverse-transcribed with random primers using the SuperScript III Reverse Transcriptase kit (Invitrogen, Carlsbad, CA, USA).

    Article Title: Simultaneous purification of DNA and RNA from microbiota in a single colonic mucosal biopsy
    Article Snippet: In the present article, we compare several tissue lysis steps and commercial purification kits, to achieve a joint RNA and DNA purification protocol for the purpose of investigating the microbiota and the microbiota-host interactions in a single colonic mucosal tissue sample. .. A further optimised tissue homogenisation and lysis protocol comprising mechanical bead beating, lysis buffer replacement and enzymatic treatment, in combination with the AllPrep DNA/RNA Mini Kit (Qiagen, Hilden, Germany) resulted in efficient and simultaneous purification of microbial and human RNA and DNA from a single mucosal colonic tissue sample. .. The present work provides a unique possibility to study RNA and DNA from the same mucosal biopsy sample, making a direct comparison between metabolically active microbes and total microbial DNA.

    Article Title: Integrating host response and unbiased microbe detection for lower respiratory tract infection diagnosis in critically ill adults
    Article Snippet: Excess TA was collected on ice, mixed 1:1 with DNA/RNA Shield (Zymo), and frozen at −80 °C. .. RNA and DNA were extracted from 300 µL of patient TA using bead-based lysis and the Allprep DNA/RNA kit (Qiagen). .. RNA was reverse transcribed to generate cDNA and used to construct sequencing libraries using the NEBNext Ultra II Library Prep Kit (New England Biolabs).

    Article Title: Simultaneous purification of DNA and RNA from microbiota in a single colonic mucosal biopsy
    Article Snippet: In the tissue lysis protocol of the AllPrep DNA/RNA Mini Kit, RNases and other proteins are efficiently denatured by β-mercaptoethanol added in the buffer RLT Plus. .. The finding of bead beating and enzymatic lysis in combination with the AllPrep DNA/RNA Mini Kit being superior in breaking the ‘hard-to-lyse’ cell walls of the Firmicutes phylum (Fig. ) resulted in development of protocol 3 in an attempt to use enzymatic lysis and bead beating and still preserve microbial and human RNA. .. Protocol 3 is a combination of protocol 2 and a modified version of the enzymatic lysis procedure for stool samples published by Franzosa and colleagues [ ].

    Homogenization:

    Article Title: Simultaneous purification of DNA and RNA from microbiota in a single colonic mucosal biopsy
    Article Snippet: In the present article, we compare several tissue lysis steps and commercial purification kits, to achieve a joint RNA and DNA purification protocol for the purpose of investigating the microbiota and the microbiota-host interactions in a single colonic mucosal tissue sample. .. A further optimised tissue homogenisation and lysis protocol comprising mechanical bead beating, lysis buffer replacement and enzymatic treatment, in combination with the AllPrep DNA/RNA Mini Kit (Qiagen, Hilden, Germany) resulted in efficient and simultaneous purification of microbial and human RNA and DNA from a single mucosal colonic tissue sample. .. The present work provides a unique possibility to study RNA and DNA from the same mucosal biopsy sample, making a direct comparison between metabolically active microbes and total microbial DNA.

    Purification:

    Article Title: Simultaneous purification of DNA and RNA from microbiota in a single colonic mucosal biopsy
    Article Snippet: In the present article, we compare several tissue lysis steps and commercial purification kits, to achieve a joint RNA and DNA purification protocol for the purpose of investigating the microbiota and the microbiota-host interactions in a single colonic mucosal tissue sample. .. A further optimised tissue homogenisation and lysis protocol comprising mechanical bead beating, lysis buffer replacement and enzymatic treatment, in combination with the AllPrep DNA/RNA Mini Kit (Qiagen, Hilden, Germany) resulted in efficient and simultaneous purification of microbial and human RNA and DNA from a single mucosal colonic tissue sample. .. The present work provides a unique possibility to study RNA and DNA from the same mucosal biopsy sample, making a direct comparison between metabolically active microbes and total microbial DNA.

    Article Title: Simultaneous purification of DNA and RNA from microbiota in a single colonic mucosal biopsy
    Article Snippet: .. Total RNA and DNA were purified from colonic mucosal tissue using the AllPrep DNA/RNA Mini Kit (Qiagen). .. Manufacturer’s instructions were followed with the exception of the lysis steps, where three tissue lysis protocols (Protocol 1, 2 and 3) were performed and evaluated (Fig. ; Additional file ).

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  • 99
    Qiagen allprep dna rna kit
    Workflow for distinguishing LRTI pathogens from commensal respiratory microbiota using an algorithmic approach. ( A ) Projection of microbial relative abundance in log reads per million reads sequenced (rpm) by <t>RNA</t> sequencing (RNA-seq) ( x axis) versus <t>DNA</t> sequencing (DNA-seq) ( y axis) for representative cases. In the LRTI +C+M group, pathogens identified by standard clinical microbiology (filled shapes) had higher overall relative abundance compared with other taxa detected by sequencing (open shapes). The largest score differential between ranked microbes (max Δrpm) was used as a threshold to identify high-scoring taxa, distinct from the other microbes based on abundance (line with arrows). Red indicates taxa represented in the reference list of established LRTI pathogens. ( B ) Receiver operating characteristic (ROC) curve demonstrating logistic regression model (LRM) performance for detecting pathogens versus commensal microbiota in both the derivation and validation cohorts. The gray ROC curve and shaded region indicate results from 1,000 rounds of training and testing on randomized sets the derivation cohort. The blue and green lines indicate predictions using leave-one-patient-out cross-validation (LOPO-CV) on the derivation and validation on the validation cohort, respectively. ( C ) Microbes predicted by the LRM to represent putative pathogens. The x axis represents combined RNA-seq and DNA-seq relative abundance, and the y axis indicates pathogen probability. The dashed line reflects the optimized probability threshold for pathogen assignment. Red filled circles: microbes predicted by LRM to represent putative LRTI pathogens that were also identified by conventional microbiologic tests. Blue filled circles: microbes predicted to represent putative LRTI pathogens by LRM only. Blue open circles: microbes identified by NGS but not predicted by the LRM to represent putative pathogens. Red open circles: microbes identified using NGS and by standard microbiologic testing but not predicted to be putative pathogens. Dark red outlined circles: microbes detected as part of a polymicrobial culture.
    Allprep Dna Rna Kit, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/allprep dna rna kit/product/Qiagen
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    allprep dna rna kit - by Bioz Stars, 2021-03
    99/100 stars
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    Image Search Results


    Workflow for distinguishing LRTI pathogens from commensal respiratory microbiota using an algorithmic approach. ( A ) Projection of microbial relative abundance in log reads per million reads sequenced (rpm) by RNA sequencing (RNA-seq) ( x axis) versus DNA sequencing (DNA-seq) ( y axis) for representative cases. In the LRTI +C+M group, pathogens identified by standard clinical microbiology (filled shapes) had higher overall relative abundance compared with other taxa detected by sequencing (open shapes). The largest score differential between ranked microbes (max Δrpm) was used as a threshold to identify high-scoring taxa, distinct from the other microbes based on abundance (line with arrows). Red indicates taxa represented in the reference list of established LRTI pathogens. ( B ) Receiver operating characteristic (ROC) curve demonstrating logistic regression model (LRM) performance for detecting pathogens versus commensal microbiota in both the derivation and validation cohorts. The gray ROC curve and shaded region indicate results from 1,000 rounds of training and testing on randomized sets the derivation cohort. The blue and green lines indicate predictions using leave-one-patient-out cross-validation (LOPO-CV) on the derivation and validation on the validation cohort, respectively. ( C ) Microbes predicted by the LRM to represent putative pathogens. The x axis represents combined RNA-seq and DNA-seq relative abundance, and the y axis indicates pathogen probability. The dashed line reflects the optimized probability threshold for pathogen assignment. Red filled circles: microbes predicted by LRM to represent putative LRTI pathogens that were also identified by conventional microbiologic tests. Blue filled circles: microbes predicted to represent putative LRTI pathogens by LRM only. Blue open circles: microbes identified by NGS but not predicted by the LRM to represent putative pathogens. Red open circles: microbes identified using NGS and by standard microbiologic testing but not predicted to be putative pathogens. Dark red outlined circles: microbes detected as part of a polymicrobial culture.

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

    Article Title: Integrating host response and unbiased microbe detection for lower respiratory tract infection diagnosis in critically ill adults

    doi: 10.1073/pnas.1809700115

    Figure Lengend Snippet: Workflow for distinguishing LRTI pathogens from commensal respiratory microbiota using an algorithmic approach. ( A ) Projection of microbial relative abundance in log reads per million reads sequenced (rpm) by RNA sequencing (RNA-seq) ( x axis) versus DNA sequencing (DNA-seq) ( y axis) for representative cases. In the LRTI +C+M group, pathogens identified by standard clinical microbiology (filled shapes) had higher overall relative abundance compared with other taxa detected by sequencing (open shapes). The largest score differential between ranked microbes (max Δrpm) was used as a threshold to identify high-scoring taxa, distinct from the other microbes based on abundance (line with arrows). Red indicates taxa represented in the reference list of established LRTI pathogens. ( B ) Receiver operating characteristic (ROC) curve demonstrating logistic regression model (LRM) performance for detecting pathogens versus commensal microbiota in both the derivation and validation cohorts. The gray ROC curve and shaded region indicate results from 1,000 rounds of training and testing on randomized sets the derivation cohort. The blue and green lines indicate predictions using leave-one-patient-out cross-validation (LOPO-CV) on the derivation and validation on the validation cohort, respectively. ( C ) Microbes predicted by the LRM to represent putative pathogens. The x axis represents combined RNA-seq and DNA-seq relative abundance, and the y axis indicates pathogen probability. The dashed line reflects the optimized probability threshold for pathogen assignment. Red filled circles: microbes predicted by LRM to represent putative LRTI pathogens that were also identified by conventional microbiologic tests. Blue filled circles: microbes predicted to represent putative LRTI pathogens by LRM only. Blue open circles: microbes identified by NGS but not predicted by the LRM to represent putative pathogens. Red open circles: microbes identified using NGS and by standard microbiologic testing but not predicted to be putative pathogens. Dark red outlined circles: microbes detected as part of a polymicrobial culture.

    Article Snippet: RNA and DNA were extracted from 300 µL of patient TA using bead-based lysis and the Allprep DNA/RNA kit (Qiagen).

    Techniques: RNA Sequencing Assay, DNA Sequencing, Sequencing, Next-Generation Sequencing

    Study overview and analysis workflow. Patients with acute respiratory failure were enrolled within 72 h of ICU admission, and TA samples were collected and underwent both RNA sequencing (RNA-seq) and shotgun DNA sequencing (DNA-seq). Post hoc clinical adjudication blinded to mNGS results identified patients with LRTI defined by clinical and microbiologic criteria (LRTI +C+M ); LRTI defined by clinical criteria only (LRTI +C ); patients with noninfectious reasons for acute respiratory failure (no-LRTI); and respiratory failure due to unknown cause (unk-LRTI). The LRTI +C+M and no-LRTI groups were divided into derivation and validation cohorts. To detect pathogens and differentiate them from a background of commensal microbiota, we developed two models: a rules-based model (RBM) and a logistic regression model (LRM). LRTI probability was next evaluated with ( i ) a pathogen metric, ( ii ) a lung microbiome diversity metric, and ( iii ) a 12-gene host transcriptional classifier. Models were then combined and optimized for LRTI rule out.

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

    Article Title: Integrating host response and unbiased microbe detection for lower respiratory tract infection diagnosis in critically ill adults

    doi: 10.1073/pnas.1809700115

    Figure Lengend Snippet: Study overview and analysis workflow. Patients with acute respiratory failure were enrolled within 72 h of ICU admission, and TA samples were collected and underwent both RNA sequencing (RNA-seq) and shotgun DNA sequencing (DNA-seq). Post hoc clinical adjudication blinded to mNGS results identified patients with LRTI defined by clinical and microbiologic criteria (LRTI +C+M ); LRTI defined by clinical criteria only (LRTI +C ); patients with noninfectious reasons for acute respiratory failure (no-LRTI); and respiratory failure due to unknown cause (unk-LRTI). The LRTI +C+M and no-LRTI groups were divided into derivation and validation cohorts. To detect pathogens and differentiate them from a background of commensal microbiota, we developed two models: a rules-based model (RBM) and a logistic regression model (LRM). LRTI probability was next evaluated with ( i ) a pathogen metric, ( ii ) a lung microbiome diversity metric, and ( iii ) a 12-gene host transcriptional classifier. Models were then combined and optimized for LRTI rule out.

    Article Snippet: RNA and DNA were extracted from 300 µL of patient TA using bead-based lysis and the Allprep DNA/RNA kit (Qiagen).

    Techniques: RNA Sequencing Assay, DNA Sequencing

    Clinical course of the London patient up to 29 months after analytical treatment interruption Upper panel shows peripheral blood CD4 count, plasma HIV-1 RNA, HIV-1 DNA, and chimerism in peripheral T cells over time. Lower panel shows amounts in DNA of CMV and EBV in plasma over time. Anti-CD52 was alemtuzumab. 3TC=lamivudine. Allo-HSCT=allogeneic haemopoietic stem-cell transplantation. cART=combination antiretroviral therapy. CMV=cytomegalovirus. CsA=ciclosporin. DTG=dolutegravir. EBV=Epstein-Barr virus. GvHD=graft versus host disease. FTC=emtricitabine. LACE=lomustine, cytarabine, cyclophosphamide, and etoposide. MTX=methotrexate. RAL=raltegravir. RPV=rilpivirine. TDF=tenofovir disoproxil fumarate.

    Journal: The Lancet. HIV

    Article Title: Evidence for HIV-1 cure after CCR5Δ32/Δ32 allogeneic haemopoietic stem-cell transplantation 30 months post analytical treatment interruption: a case report

    doi: 10.1016/S2352-3018(20)30069-2

    Figure Lengend Snippet: Clinical course of the London patient up to 29 months after analytical treatment interruption Upper panel shows peripheral blood CD4 count, plasma HIV-1 RNA, HIV-1 DNA, and chimerism in peripheral T cells over time. Lower panel shows amounts in DNA of CMV and EBV in plasma over time. Anti-CD52 was alemtuzumab. 3TC=lamivudine. Allo-HSCT=allogeneic haemopoietic stem-cell transplantation. cART=combination antiretroviral therapy. CMV=cytomegalovirus. CsA=ciclosporin. DTG=dolutegravir. EBV=Epstein-Barr virus. GvHD=graft versus host disease. FTC=emtricitabine. LACE=lomustine, cytarabine, cyclophosphamide, and etoposide. MTX=methotrexate. RAL=raltegravir. RPV=rilpivirine. TDF=tenofovir disoproxil fumarate.

    Article Snippet: DNA was extracted using an Qiagen AllPrep DNA/RNA Mini kit.

    Techniques: Transplantation Assay

    Phylogenetic relationship and relative abundance of SAR11 OTUs. Only SAR11 OTUs representing > 0.2% of total SAR11 OTU sequences are included. Bubble size is scaled to sequence abundance of OTUs relative to total SAR11 communities. DNA and RNA is 16S rRNA gene copies and 16S rRNA, respectively. High and low diversity sub-groups were empirically defined; see Figure 1 and text for further details.

    Journal: The ISME Journal

    Article Title: Seasonal dynamics of active SAR11 ecotypes in the oligotrophic Northwest Mediterranean Sea

    doi: 10.1038/ismej.2014.129

    Figure Lengend Snippet: Phylogenetic relationship and relative abundance of SAR11 OTUs. Only SAR11 OTUs representing > 0.2% of total SAR11 OTU sequences are included. Bubble size is scaled to sequence abundance of OTUs relative to total SAR11 communities. DNA and RNA is 16S rRNA gene copies and 16S rRNA, respectively. High and low diversity sub-groups were empirically defined; see Figure 1 and text for further details.

    Article Snippet: A combination of mechanic (freeze-thaw) and enzymatic cell lysis techniques were applied directly to Sterivex cartridges, followed by extraction using an AllPrep DNA/RNA kit (Qiagen, Venlo, Netherlands).

    Techniques: Sequencing

    Leucine incorporation and RNA/DNA ratios in the SAR11 clade . The ratio of ribosomal RNA (rRNA) to ribosomal RNA gene copies (rDNA) were calculated by summing the sequences for all SAR11 clusters. SAR11-active cells were calculated as the percentage of total SAR11 cells assimilating tritiated leucine.

    Journal: The ISME Journal

    Article Title: Seasonal dynamics of active SAR11 ecotypes in the oligotrophic Northwest Mediterranean Sea

    doi: 10.1038/ismej.2014.129

    Figure Lengend Snippet: Leucine incorporation and RNA/DNA ratios in the SAR11 clade . The ratio of ribosomal RNA (rRNA) to ribosomal RNA gene copies (rDNA) were calculated by summing the sequences for all SAR11 clusters. SAR11-active cells were calculated as the percentage of total SAR11 cells assimilating tritiated leucine.

    Article Snippet: A combination of mechanic (freeze-thaw) and enzymatic cell lysis techniques were applied directly to Sterivex cartridges, followed by extraction using an AllPrep DNA/RNA kit (Qiagen, Venlo, Netherlands).

    Techniques:

    Exon splicing analysis of the BRCA1 c.5152+6T > C variant of patient PT51. A, Schematic view of variant c.5152+6T > C localization in the BRCA1 gene. PCR primer alignment is indicated with the red and blue bars. Sequencing analysis for genomic DNA is presented below. B, RT‐PCR of lymphocyte‐derived RNA. Predicted scheme of mRNA transcript in control or patient samples (upper right panel). Agarose gel (2%) electrophoresis; lane 1: control sample; lane 2: patient sample. Two PCR products were detected in the patient sample (upper middle panel). Chromatogram sequences of the control and abnormal transcripts. Vertical line in the chromatogram indicates the exonic junction in transcripts. Exon 18 (78 bp) skipping between exon 17 and exon 19 was identified (upper left panel). Functional domains of BRCA1 and sequence alignment of the BRCA1 abnormal transcript (lower panel). Amino acid sequences of the splice variant (c.5152+6T > C) were aligned using a reference sequence (NP_009225.1) via NCBI BLAST ( https://blast.ncbi.nlm.nih.gov/Blast.cgi ). BRCA1 c.5152+6T > C was identified to encode a BRCA1 protein with an in‐frame deletion (26 amino acids) in the BRCA1 C‐terminal (BRCT) domain; this may affect the function of the BRCA1 BRCT domain. The red line indicates the location of the in‐frame deletion residues. C, Pedigree of patient PT51

    Journal: Cancer Science

    Article Title: Exon splicing analysis of intronic variants in multigene cancer panel testing for hereditary breast/ovarian cancer, et al. Exon splicing analysis of intronic variants in multigene cancer panel testing for hereditary breast/ovarian cancer

    doi: 10.1111/cas.14600

    Figure Lengend Snippet: Exon splicing analysis of the BRCA1 c.5152+6T > C variant of patient PT51. A, Schematic view of variant c.5152+6T > C localization in the BRCA1 gene. PCR primer alignment is indicated with the red and blue bars. Sequencing analysis for genomic DNA is presented below. B, RT‐PCR of lymphocyte‐derived RNA. Predicted scheme of mRNA transcript in control or patient samples (upper right panel). Agarose gel (2%) electrophoresis; lane 1: control sample; lane 2: patient sample. Two PCR products were detected in the patient sample (upper middle panel). Chromatogram sequences of the control and abnormal transcripts. Vertical line in the chromatogram indicates the exonic junction in transcripts. Exon 18 (78 bp) skipping between exon 17 and exon 19 was identified (upper left panel). Functional domains of BRCA1 and sequence alignment of the BRCA1 abnormal transcript (lower panel). Amino acid sequences of the splice variant (c.5152+6T > C) were aligned using a reference sequence (NP_009225.1) via NCBI BLAST ( https://blast.ncbi.nlm.nih.gov/Blast.cgi ). BRCA1 c.5152+6T > C was identified to encode a BRCA1 protein with an in‐frame deletion (26 amino acids) in the BRCA1 C‐terminal (BRCT) domain; this may affect the function of the BRCA1 BRCT domain. The red line indicates the location of the in‐frame deletion residues. C, Pedigree of patient PT51

    Article Snippet: Total RNA was extracted from peripheral blood lymphocytes using the NucleoSpin RNA Blood Kit (Macherey‐Nagel) or from normal tissues using the AllPrep DNA/RNA Mini Kit (Qiagen) according to manufacturer's instructions.

    Techniques: Variant Assay, Polymerase Chain Reaction, Sequencing, Reverse Transcription Polymerase Chain Reaction, Derivative Assay, Agarose Gel Electrophoresis, Electrophoresis, Functional Assay