pcr analysis  (Qiagen)

 
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    QIAamp MinElute Virus Spin Kit
    Description:
    For simultaneous purification of viral DNA and RNA from plasma serum and cell free body fluids Kit contents Qiagen QIAamp MinElute Virus Spin Kit 50 preps 200L Sample 20 to 150L Elution Volume Serum Plasma Sample Viral DNA and RNA Purification Silica Technology Manual Processing MinElute Columns Format For Simultaneous Purification of Viral DNA and RNA from Plasma Serum and Cell free Body Fluids Ideal for PCR Real time PCR Application Includes 50 QIAamp MinElute Columns Qiagen Protease Carrier RNA Buffers 2mL Collection Tubes Benefits Rapid purification of high quality viral DNA and RNA No organic extraction or alcohol precipitation Consistent high yields Complete removal of contaminants and inhibito
    Catalog Number:
    57704
    Price:
    278
    Category:
    QIAamp MinElute Virus Spin Kit
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    Structured Review

    Qiagen pcr analysis
    QIAamp MinElute Virus Spin Kit
    For simultaneous purification of viral DNA and RNA from plasma serum and cell free body fluids Kit contents Qiagen QIAamp MinElute Virus Spin Kit 50 preps 200L Sample 20 to 150L Elution Volume Serum Plasma Sample Viral DNA and RNA Purification Silica Technology Manual Processing MinElute Columns Format For Simultaneous Purification of Viral DNA and RNA from Plasma Serum and Cell free Body Fluids Ideal for PCR Real time PCR Application Includes 50 QIAamp MinElute Columns Qiagen Protease Carrier RNA Buffers 2mL Collection Tubes Benefits Rapid purification of high quality viral DNA and RNA No organic extraction or alcohol precipitation Consistent high yields Complete removal of contaminants and inhibito
    https://www.bioz.com/result/pcr analysis/product/Qiagen
    Average 94 stars, based on 7014 article reviews
    Price from $9.99 to $1999.99
    pcr analysis - by Bioz Stars, 2020-07
    94/100 stars

    Images

    1) Product Images from "Bioengineered Viral Platform for Intramuscular Passive Vaccine Delivery to Human Skeletal Muscle"

    Article Title: Bioengineered Viral Platform for Intramuscular Passive Vaccine Delivery to Human Skeletal Muscle

    Journal: Molecular Therapy. Methods & Clinical Development

    doi: 10.1016/j.omtm.2018.06.001

    Directed Evolution of AAV Capsids by DNA Shuffling and Selection Screening on Primary Human Skeletal Muscle Stem Cells and Myotubes (A) AAV capsid genes from ten parental serotypes (1, 2, 3b, 4, 5, 6, 8, 9_hu14, avian, and bovine) were PCR-amplified, fragmented with DNaseI digestion, and then randomly reassembled through self-priming PCR. Resultant shuffled Cap genes were cloned back into a replication-competent AAV production plasmid via flanking SwaI/NsiI sites downstream of AAV2 Rep dot blot titered, and used for selection. (B) Sequential gating scheme Panels [P]1–P4) to isolate CD31 − CD34 − CD45 − ITGB1 + EGFR + primary human skeletal muscle stem cells from surgical specimens. Numbers indicate percentage of total events falling within each gate. (C) Phase contrast and immunofluorescence PAX7 (pink) and DAPI (blue) nuclear staining of purified primary human muscle stem cell cultures. Scale bar, 100 μm. (D) Diagram illustrating the two selection screens with replicating AAV capsid libraries from (A) that was performed on both pooled primary human skeletal muscle stem cells and human myotubes. (E) Semiquantitative AAV Cap PCR (2.2-Kb product) was performed at each round of each selection screen to demonstrate active replication of AAV library genomes throughout each round of the screen.
    Figure Legend Snippet: Directed Evolution of AAV Capsids by DNA Shuffling and Selection Screening on Primary Human Skeletal Muscle Stem Cells and Myotubes (A) AAV capsid genes from ten parental serotypes (1, 2, 3b, 4, 5, 6, 8, 9_hu14, avian, and bovine) were PCR-amplified, fragmented with DNaseI digestion, and then randomly reassembled through self-priming PCR. Resultant shuffled Cap genes were cloned back into a replication-competent AAV production plasmid via flanking SwaI/NsiI sites downstream of AAV2 Rep dot blot titered, and used for selection. (B) Sequential gating scheme Panels [P]1–P4) to isolate CD31 − CD34 − CD45 − ITGB1 + EGFR + primary human skeletal muscle stem cells from surgical specimens. Numbers indicate percentage of total events falling within each gate. (C) Phase contrast and immunofluorescence PAX7 (pink) and DAPI (blue) nuclear staining of purified primary human muscle stem cell cultures. Scale bar, 100 μm. (D) Diagram illustrating the two selection screens with replicating AAV capsid libraries from (A) that was performed on both pooled primary human skeletal muscle stem cells and human myotubes. (E) Semiquantitative AAV Cap PCR (2.2-Kb product) was performed at each round of each selection screen to demonstrate active replication of AAV library genomes throughout each round of the screen.

    Techniques Used: Selection, Polymerase Chain Reaction, Amplification, Clone Assay, Plasmid Preparation, Dot Blot, Immunofluorescence, Staining, Purification

    2) Product Images from "Bioengineered Viral Platform for Intramuscular Passive Vaccine Delivery to Human Skeletal Muscle"

    Article Title: Bioengineered Viral Platform for Intramuscular Passive Vaccine Delivery to Human Skeletal Muscle

    Journal: Molecular Therapy. Methods & Clinical Development

    doi: 10.1016/j.omtm.2018.06.001

    Directed Evolution of AAV Capsids by DNA Shuffling and Selection Screening on Primary Human Skeletal Muscle Stem Cells and Myotubes (A) AAV capsid genes from ten parental serotypes (1, 2, 3b, 4, 5, 6, 8, 9_hu14, avian, and bovine) were PCR-amplified, fragmented with DNaseI digestion, and then randomly reassembled through self-priming PCR. Resultant shuffled Cap genes were cloned back into a replication-competent AAV production plasmid via flanking SwaI/NsiI sites downstream of AAV2 Rep dot blot titered, and used for selection. (B) Sequential gating scheme Panels [P]1–P4) to isolate CD31 − CD34 − CD45 − ITGB1 + EGFR + primary human skeletal muscle stem cells from surgical specimens. Numbers indicate percentage of total events falling within each gate. (C) Phase contrast and immunofluorescence PAX7 (pink) and DAPI (blue) nuclear staining of purified primary human muscle stem cell cultures. Scale bar, 100 μm. (D) Diagram illustrating the two selection screens with replicating AAV capsid libraries from (A) that was performed on both pooled primary human skeletal muscle stem cells and human myotubes. (E) Semiquantitative AAV Cap PCR (2.2-Kb product) was performed at each round of each selection screen to demonstrate active replication of AAV library genomes throughout each round of the screen.
    Figure Legend Snippet: Directed Evolution of AAV Capsids by DNA Shuffling and Selection Screening on Primary Human Skeletal Muscle Stem Cells and Myotubes (A) AAV capsid genes from ten parental serotypes (1, 2, 3b, 4, 5, 6, 8, 9_hu14, avian, and bovine) were PCR-amplified, fragmented with DNaseI digestion, and then randomly reassembled through self-priming PCR. Resultant shuffled Cap genes were cloned back into a replication-competent AAV production plasmid via flanking SwaI/NsiI sites downstream of AAV2 Rep dot blot titered, and used for selection. (B) Sequential gating scheme Panels [P]1–P4) to isolate CD31 − CD34 − CD45 − ITGB1 + EGFR + primary human skeletal muscle stem cells from surgical specimens. Numbers indicate percentage of total events falling within each gate. (C) Phase contrast and immunofluorescence PAX7 (pink) and DAPI (blue) nuclear staining of purified primary human muscle stem cell cultures. Scale bar, 100 μm. (D) Diagram illustrating the two selection screens with replicating AAV capsid libraries from (A) that was performed on both pooled primary human skeletal muscle stem cells and human myotubes. (E) Semiquantitative AAV Cap PCR (2.2-Kb product) was performed at each round of each selection screen to demonstrate active replication of AAV library genomes throughout each round of the screen.

    Techniques Used: Selection, Polymerase Chain Reaction, Amplification, Clone Assay, Plasmid Preparation, Dot Blot, Immunofluorescence, Staining, Purification

    3) Product Images from "Selection and evaluation of an efficient method for the recovery of viral nucleic acids from complex biologicals"

    Article Title: Selection and evaluation of an efficient method for the recovery of viral nucleic acids from complex biologicals

    Journal: NPJ Vaccines

    doi: 10.1038/s41541-018-0067-3

    Efficiency of separate extraction of DNA and RNA when compared to total nucleic acid extraction using QIAamp® MinElute® Virus Spin kit. DNA and RNA were extracted separately and compared to the total nucleic acid extracted using the QIAamp® MinElute® Virus Spin kit, using qPCR. RNA extraction using the Qiagen RNeasy® Mini kit did not enhance the detection of the two single-stranded RNA viruses (FeLV and RSV), whereas the RNase A digestion added to the Wako DNA Extractor® kit greatly enriched for double-stranded nucleic acid (EBV and Reo3)
    Figure Legend Snippet: Efficiency of separate extraction of DNA and RNA when compared to total nucleic acid extraction using QIAamp® MinElute® Virus Spin kit. DNA and RNA were extracted separately and compared to the total nucleic acid extracted using the QIAamp® MinElute® Virus Spin kit, using qPCR. RNA extraction using the Qiagen RNeasy® Mini kit did not enhance the detection of the two single-stranded RNA viruses (FeLV and RSV), whereas the RNase A digestion added to the Wako DNA Extractor® kit greatly enriched for double-stranded nucleic acid (EBV and Reo3)

    Techniques Used: Real-time Polymerase Chain Reaction, RNA Extraction

    4) Product Images from "Performance of the digene LQ, RH and PS HPVs genotyping systems on clinical samples and comparison with HC2 and PCR-based Linear Array"

    Article Title: Performance of the digene LQ, RH and PS HPVs genotyping systems on clinical samples and comparison with HC2 and PCR-based Linear Array

    Journal: Infectious Agents and Cancer

    doi: 10.1186/1750-9378-6-23

    Type specific concordance between LA and LQ, RH and PS techniques (N = 305) . The genotyping results for each of the tested techniques vs. LA are presented in a color-coded way. Concordances LA/LQ are presented in black, LA/RH in red and LA/PS in blue.
    Figure Legend Snippet: Type specific concordance between LA and LQ, RH and PS techniques (N = 305) . The genotyping results for each of the tested techniques vs. LA are presented in a color-coded way. Concordances LA/LQ are presented in black, LA/RH in red and LA/PS in blue.

    Techniques Used:

    5) Product Images from "Selection and evaluation of an efficient method for the recovery of viral nucleic acids from complex biologicals"

    Article Title: Selection and evaluation of an efficient method for the recovery of viral nucleic acids from complex biologicals

    Journal: NPJ Vaccines

    doi: 10.1038/s41541-018-0067-3

    Efficiency of separate extraction of DNA and RNA when compared to total nucleic acid extraction using QIAamp® MinElute® Virus Spin kit. DNA and RNA were extracted separately and compared to the total nucleic acid extracted using the QIAamp® MinElute® Virus Spin kit, using qPCR. RNA extraction using the Qiagen RNeasy® Mini kit did not enhance the detection of the two single-stranded RNA viruses (FeLV and RSV), whereas the RNase A digestion added to the Wako DNA Extractor® kit greatly enriched for double-stranded nucleic acid (EBV and Reo3)
    Figure Legend Snippet: Efficiency of separate extraction of DNA and RNA when compared to total nucleic acid extraction using QIAamp® MinElute® Virus Spin kit. DNA and RNA were extracted separately and compared to the total nucleic acid extracted using the QIAamp® MinElute® Virus Spin kit, using qPCR. RNA extraction using the Qiagen RNeasy® Mini kit did not enhance the detection of the two single-stranded RNA viruses (FeLV and RSV), whereas the RNase A digestion added to the Wako DNA Extractor® kit greatly enriched for double-stranded nucleic acid (EBV and Reo3)

    Techniques Used: Real-time Polymerase Chain Reaction, RNA Extraction

    6) Product Images from "Selection and evaluation of an efficient method for the recovery of viral nucleic acids from complex biologicals"

    Article Title: Selection and evaluation of an efficient method for the recovery of viral nucleic acids from complex biologicals

    Journal: NPJ Vaccines

    doi: 10.1038/s41541-018-0067-3

    Efficiency of separate extraction of DNA and RNA when compared to total nucleic acid extraction using QIAamp® MinElute® Virus Spin kit. DNA and RNA were extracted separately and compared to the total nucleic acid extracted using the QIAamp® MinElute® Virus Spin kit, using qPCR. RNA extraction using the Qiagen RNeasy® Mini kit did not enhance the detection of the two single-stranded RNA viruses (FeLV and RSV), whereas the RNase A digestion added to the Wako DNA Extractor® kit greatly enriched for double-stranded nucleic acid (EBV and Reo3)
    Figure Legend Snippet: Efficiency of separate extraction of DNA and RNA when compared to total nucleic acid extraction using QIAamp® MinElute® Virus Spin kit. DNA and RNA were extracted separately and compared to the total nucleic acid extracted using the QIAamp® MinElute® Virus Spin kit, using qPCR. RNA extraction using the Qiagen RNeasy® Mini kit did not enhance the detection of the two single-stranded RNA viruses (FeLV and RSV), whereas the RNase A digestion added to the Wako DNA Extractor® kit greatly enriched for double-stranded nucleic acid (EBV and Reo3)

    Techniques Used: Real-time Polymerase Chain Reaction, RNA Extraction

    7) Product Images from "Bioengineered Viral Platform for Intramuscular Passive Vaccine Delivery to Human Skeletal Muscle"

    Article Title: Bioengineered Viral Platform for Intramuscular Passive Vaccine Delivery to Human Skeletal Muscle

    Journal: Molecular Therapy. Methods & Clinical Development

    doi: 10.1016/j.omtm.2018.06.001

    Directed Evolution of AAV Capsids by DNA Shuffling and Selection Screening on Primary Human Skeletal Muscle Stem Cells and Myotubes (A) AAV capsid genes from ten parental serotypes (1, 2, 3b, 4, 5, 6, 8, 9_hu14, avian, and bovine) were PCR-amplified, fragmented with DNaseI digestion, and then randomly reassembled through self-priming PCR. Resultant shuffled Cap genes were cloned back into a replication-competent AAV production plasmid via flanking SwaI/NsiI sites downstream of AAV2 Rep . The resultant library production plasmid contained AAV2 ITRs and a modified AAV2 3′ UTR. The AAV library was packaged using standard production protocols as done previously, 41 dot blot titered, and used for selection. (B) Sequential gating scheme Panels [P]1–P4) to isolate CD31 − CD34 − CD45 − ITGB1 + EGFR + primary human skeletal muscle stem cells from surgical specimens. Numbers indicate percentage of total events falling within each gate. (C) Phase contrast and immunofluorescence PAX7 (pink) and DAPI (blue) nuclear staining of purified primary human muscle stem cell cultures. Scale bar, 100 μm. (D) Diagram illustrating the two selection screens with replicating AAV capsid libraries from (A) that was performed on both pooled primary human skeletal muscle stem cells and human myotubes. (E) Semiquantitative AAV Cap PCR (2.2-Kb product) was performed at each round of each selection screen to demonstrate active replication of AAV library genomes throughout each round of the screen.
    Figure Legend Snippet: Directed Evolution of AAV Capsids by DNA Shuffling and Selection Screening on Primary Human Skeletal Muscle Stem Cells and Myotubes (A) AAV capsid genes from ten parental serotypes (1, 2, 3b, 4, 5, 6, 8, 9_hu14, avian, and bovine) were PCR-amplified, fragmented with DNaseI digestion, and then randomly reassembled through self-priming PCR. Resultant shuffled Cap genes were cloned back into a replication-competent AAV production plasmid via flanking SwaI/NsiI sites downstream of AAV2 Rep . The resultant library production plasmid contained AAV2 ITRs and a modified AAV2 3′ UTR. The AAV library was packaged using standard production protocols as done previously, 41 dot blot titered, and used for selection. (B) Sequential gating scheme Panels [P]1–P4) to isolate CD31 − CD34 − CD45 − ITGB1 + EGFR + primary human skeletal muscle stem cells from surgical specimens. Numbers indicate percentage of total events falling within each gate. (C) Phase contrast and immunofluorescence PAX7 (pink) and DAPI (blue) nuclear staining of purified primary human muscle stem cell cultures. Scale bar, 100 μm. (D) Diagram illustrating the two selection screens with replicating AAV capsid libraries from (A) that was performed on both pooled primary human skeletal muscle stem cells and human myotubes. (E) Semiquantitative AAV Cap PCR (2.2-Kb product) was performed at each round of each selection screen to demonstrate active replication of AAV library genomes throughout each round of the screen.

    Techniques Used: Selection, Polymerase Chain Reaction, Amplification, Clone Assay, Plasmid Preparation, Modification, Dot Blot, Immunofluorescence, Staining, Purification

    8) Product Images from "An RNA-based system to study hepatitis B virus replication and select drug-resistance mutations"

    Article Title: An RNA-based system to study hepatitis B virus replication and select drug-resistance mutations

    Journal: bioRxiv

    doi: 10.1101/787630

    RNA launch method can be used to test drug efficiency and identify drug-resistance mutations. ( A ) HBV reverse transcriptase inhibitor dose response six days post transfection with HBV genotype A pgRNA. Secreted HBsAg was quantified by CLIA and normalized to untreated controls. Data plotted are n=3 ( B ) Sequencing extracellular HBV DNA from cells transfected with HBV pgRNA in the presence of 400 μM LAM identifies the two most common amino acid substitutions, M204V and M204I, found in LAM-resistant patients. Threshold = Bonferroni correction, alpha = 0.05.
    Figure Legend Snippet: RNA launch method can be used to test drug efficiency and identify drug-resistance mutations. ( A ) HBV reverse transcriptase inhibitor dose response six days post transfection with HBV genotype A pgRNA. Secreted HBsAg was quantified by CLIA and normalized to untreated controls. Data plotted are n=3 ( B ) Sequencing extracellular HBV DNA from cells transfected with HBV pgRNA in the presence of 400 μM LAM identifies the two most common amino acid substitutions, M204V and M204I, found in LAM-resistant patients. Threshold = Bonferroni correction, alpha = 0.05.

    Techniques Used: Transfection, Sequencing, Laser Capture Microdissection

    Selecting drug-resistant viral variants from in vitro -transcribed HBV pgRNA. A ) Schematic describes a method to select and enrich mutations that confer resistance to HBV antivirals. From left, T7 bacteriophage RNA polymerase generates a diverse population of HBV pgRNA variants. These RNAs, transfected into cells in the presence of anti-HBV drugs, are subject to selection. HBV DNA, either from cell lysates, in secreted virions, or both, are collected, amplified by PCR, and sequenced with next generation sequencing to identify rare drug-resistant variants. Depending on the selective pressure, reverse-transcribed DNA in the cell, encapsidated DNA in the supernatant, or both, can be PCR amplified and sequenced to enrich for HBV variants. ( B ) Error rate of T7 bacteriophage RNA polymerase determined by CirSeq using in vitro -transcribed HBV pgRNA. ( C ) 1% agarose gel of HBV DNA amplified two days post transfection from supernatants of cells untreated or treated with 40 or 400 μM LAM.
    Figure Legend Snippet: Selecting drug-resistant viral variants from in vitro -transcribed HBV pgRNA. A ) Schematic describes a method to select and enrich mutations that confer resistance to HBV antivirals. From left, T7 bacteriophage RNA polymerase generates a diverse population of HBV pgRNA variants. These RNAs, transfected into cells in the presence of anti-HBV drugs, are subject to selection. HBV DNA, either from cell lysates, in secreted virions, or both, are collected, amplified by PCR, and sequenced with next generation sequencing to identify rare drug-resistant variants. Depending on the selective pressure, reverse-transcribed DNA in the cell, encapsidated DNA in the supernatant, or both, can be PCR amplified and sequenced to enrich for HBV variants. ( B ) Error rate of T7 bacteriophage RNA polymerase determined by CirSeq using in vitro -transcribed HBV pgRNA. ( C ) 1% agarose gel of HBV DNA amplified two days post transfection from supernatants of cells untreated or treated with 40 or 400 μM LAM.

    Techniques Used: In Vitro, Transfection, Selection, Amplification, Polymerase Chain Reaction, Next-Generation Sequencing, Agarose Gel Electrophoresis, Laser Capture Microdissection

    HBV lifecycle and mRNA transcripts. ( A ) HBV lifecycle. HBV (1) enters hepatocytes via heparin sulfate proteoglycans (HSPGs) and sodium taurocholate cotransporting polypeptide (NTCP). In its encapsidated form, the genome is partially double-stranded, and upon entry to the nucleus, is converted from relaxed circular (rc) DNA to covalently closed circular (ccc) DNA. cccDNA is (2) transcribed by cellular Pol II to produce all viral mRNAs, which are then exported to the cytoplasm and (3) translated into viral proteins. Of these mRNAs, pgRNA, serves as the template for reverse transcription and also encodes the core and polymerase proteins. The encapsidated pgRNA (4) is reverse transcribed (5) by HBV polymerase and can either (6a) associate with the viral surface antigens, HBsAg, and be exported from the cell, or (6b) “recycle” to the nucleus and replenish cccDNA. ( B ) HBV genome and mRNA transcripts. Left, the highly compact, 3.2 kb double stranded HBV DNA genome encodes multiple overlapping open reading frames (ORFs). Colored boxes indicate ORFs and arrows indicate Pol II transcriptional start sites. Right, the five HBV mRNAs. All transcripts terminate at the same cleavage and polyA addition signal.
    Figure Legend Snippet: HBV lifecycle and mRNA transcripts. ( A ) HBV lifecycle. HBV (1) enters hepatocytes via heparin sulfate proteoglycans (HSPGs) and sodium taurocholate cotransporting polypeptide (NTCP). In its encapsidated form, the genome is partially double-stranded, and upon entry to the nucleus, is converted from relaxed circular (rc) DNA to covalently closed circular (ccc) DNA. cccDNA is (2) transcribed by cellular Pol II to produce all viral mRNAs, which are then exported to the cytoplasm and (3) translated into viral proteins. Of these mRNAs, pgRNA, serves as the template for reverse transcription and also encodes the core and polymerase proteins. The encapsidated pgRNA (4) is reverse transcribed (5) by HBV polymerase and can either (6a) associate with the viral surface antigens, HBsAg, and be exported from the cell, or (6b) “recycle” to the nucleus and replenish cccDNA. ( B ) HBV genome and mRNA transcripts. Left, the highly compact, 3.2 kb double stranded HBV DNA genome encodes multiple overlapping open reading frames (ORFs). Colored boxes indicate ORFs and arrows indicate Pol II transcriptional start sites. Right, the five HBV mRNAs. All transcripts terminate at the same cleavage and polyA addition signal.

    Techniques Used: Countercurrent Chromatography

    Existing cell culture systems to study HBV. ( A ) A plasmid-based method to initiate HBV replication in cell culture. HepG2 cells were transfected with a plasmid containing over-length (1.3x) HBV genotype A. Anti-HBcAg immunofluorescence indicates that approximately 70-90% of transfected cells stain positive for HBcAg. qPCR detects high HBV DNA copies; however, the signal emanates largely from the transfected plasmid since both WT and Pol -- (YMHD) sequences yield similar DNA copy number. Likewise, HBsAg detected in cells by immunofluorescence or in supernatants by CLIA is produced from the transfected plasmid. ( B ) HepDE19 cell line with inducible pgRNA produced from an integrated copy of genotype D HBV. Cells were cultured with (uninduced) and without (induced) tetracycline for five days then either fixed and stained or harvested for DNA extraction. HBcAg immunofluorescence indicates that pgRNA induction increases the percentage of HBcAg-positive cells and qPCR shows a greater than 100-fold increase in HBV DNA copies. (The ∼10 6 copies of HBV DNA detected in the uninduced condition is likely from HBV DNA integrated in the cellular genome.) Despite clear differences in HBcAg and HBV DNA with and without induction, the HBsAg protein is an unreliable measure of HBV replication since the majority of signal originates from the integrated HBV genome and is independent of replication.
    Figure Legend Snippet: Existing cell culture systems to study HBV. ( A ) A plasmid-based method to initiate HBV replication in cell culture. HepG2 cells were transfected with a plasmid containing over-length (1.3x) HBV genotype A. Anti-HBcAg immunofluorescence indicates that approximately 70-90% of transfected cells stain positive for HBcAg. qPCR detects high HBV DNA copies; however, the signal emanates largely from the transfected plasmid since both WT and Pol -- (YMHD) sequences yield similar DNA copy number. Likewise, HBsAg detected in cells by immunofluorescence or in supernatants by CLIA is produced from the transfected plasmid. ( B ) HepDE19 cell line with inducible pgRNA produced from an integrated copy of genotype D HBV. Cells were cultured with (uninduced) and without (induced) tetracycline for five days then either fixed and stained or harvested for DNA extraction. HBcAg immunofluorescence indicates that pgRNA induction increases the percentage of HBcAg-positive cells and qPCR shows a greater than 100-fold increase in HBV DNA copies. (The ∼10 6 copies of HBV DNA detected in the uninduced condition is likely from HBV DNA integrated in the cellular genome.) Despite clear differences in HBcAg and HBV DNA with and without induction, the HBsAg protein is an unreliable measure of HBV replication since the majority of signal originates from the integrated HBV genome and is independent of replication.

    Techniques Used: Cell Culture, Plasmid Preparation, Transfection, Immunofluorescence, Staining, Real-time Polymerase Chain Reaction, Produced, DNA Extraction

    Pan-genotype HBV replication from in vitro -transcribed HBV pgRNA. ( A ) Unrooted phylogenetic tree of HBV genotypes A-H. Tree was generated using SeaView ( 22 ). Scale bar indicates nucleotide substitutions per site. ( B ) qPCR of intracellular HBV DNA for HBV genotypes A-H two days post transfection with and without 10 μM ETV. ( C ) CLIA detects secreted HBsAg from HBV genotypes A-H six days post transfection with and without 10 μM ETV. ETV, entecavir; copies/well (6-well plate); N.D., not detected; LOQ, limit of quantification.
    Figure Legend Snippet: Pan-genotype HBV replication from in vitro -transcribed HBV pgRNA. ( A ) Unrooted phylogenetic tree of HBV genotypes A-H. Tree was generated using SeaView ( 22 ). Scale bar indicates nucleotide substitutions per site. ( B ) qPCR of intracellular HBV DNA for HBV genotypes A-H two days post transfection with and without 10 μM ETV. ( C ) CLIA detects secreted HBsAg from HBV genotypes A-H six days post transfection with and without 10 μM ETV. ETV, entecavir; copies/well (6-well plate); N.D., not detected; LOQ, limit of quantification.

    Techniques Used: In Vitro, Generated, Real-time Polymerase Chain Reaction, Transfection

    The RNA launch system. From left to right, the figures shows that in vitro -transcribed pgRNA is (I) translated (HBcAg is detected by immunofluorescence in ∼80% of Huh7.5-NTCP cells), (II) reverse transcribed (qPCR detects HBV DNA and the signal is HBV polymerase-dependent, as a catalytic site mutation (Pol -- , YMHD) decreases the DNA signal by 10,000-fold), (III) is transported to the nucleus to form cccDNA (Southern blot confirms cccDNA is produced; see below for lane description), and (IV) cccDNA is transcribed to produce viral mRNA and protein (shown by HBsAg chemiluminescent immunoassay (CLIA)). HBcAg, HBV core antigen; HBsAg, HBV S antigen; WT, wildtype; LOQ, limit of quantification; IU, international units; N.D., not detected; copies/well, one well of 6-well plate; kb, kilobase; Southern blot: Lane 1, 1-kb ladder; Lane 2, WT HBV DNA; Lane 3, WT HBV DNA heated to 85°C; Lane 4, WT HBV DNA heated to 85°C then linearized with EcoRI; Lane 5, Pol -- HBV DNA; RC/L, relaxed circular/linear DNA; CCC, covalently closed circular DNA; SS, single stranded DNA.
    Figure Legend Snippet: The RNA launch system. From left to right, the figures shows that in vitro -transcribed pgRNA is (I) translated (HBcAg is detected by immunofluorescence in ∼80% of Huh7.5-NTCP cells), (II) reverse transcribed (qPCR detects HBV DNA and the signal is HBV polymerase-dependent, as a catalytic site mutation (Pol -- , YMHD) decreases the DNA signal by 10,000-fold), (III) is transported to the nucleus to form cccDNA (Southern blot confirms cccDNA is produced; see below for lane description), and (IV) cccDNA is transcribed to produce viral mRNA and protein (shown by HBsAg chemiluminescent immunoassay (CLIA)). HBcAg, HBV core antigen; HBsAg, HBV S antigen; WT, wildtype; LOQ, limit of quantification; IU, international units; N.D., not detected; copies/well, one well of 6-well plate; kb, kilobase; Southern blot: Lane 1, 1-kb ladder; Lane 2, WT HBV DNA; Lane 3, WT HBV DNA heated to 85°C; Lane 4, WT HBV DNA heated to 85°C then linearized with EcoRI; Lane 5, Pol -- HBV DNA; RC/L, relaxed circular/linear DNA; CCC, covalently closed circular DNA; SS, single stranded DNA.

    Techniques Used: In Vitro, Immunofluorescence, Real-time Polymerase Chain Reaction, Mutagenesis, Southern Blot, Produced, Countercurrent Chromatography

    9) Product Images from "Bioengineered Viral Platform for Intramuscular Passive Vaccine Delivery to Human Skeletal Muscle"

    Article Title: Bioengineered Viral Platform for Intramuscular Passive Vaccine Delivery to Human Skeletal Muscle

    Journal: Molecular Therapy. Methods & Clinical Development

    doi: 10.1016/j.omtm.2018.06.001

    Directed Evolution of AAV Capsids by DNA Shuffling and Selection Screening on Primary Human Skeletal Muscle Stem Cells and Myotubes (A) AAV capsid genes from ten parental serotypes (1, 2, 3b, 4, 5, 6, 8, 9_hu14, avian, and bovine) were PCR-amplified, fragmented with DNaseI digestion, and then randomly reassembled through self-priming PCR. Resultant shuffled Cap genes were cloned back into a replication-competent AAV production plasmid via flanking SwaI/NsiI sites downstream of AAV2 Rep . The resultant library production plasmid contained AAV2 ITRs and a modified AAV2 3′ UTR. The AAV library was packaged using standard production protocols as done previously, 41 dot blot titered, and used for selection. (B) Sequential gating scheme Panels [P]1–P4) to isolate CD31 − CD34 − CD45 − ITGB1 + EGFR + primary human skeletal muscle stem cells from surgical specimens. Numbers indicate percentage of total events falling within each gate. (C) Phase contrast and immunofluorescence PAX7 (pink) and DAPI (blue) nuclear staining of purified primary human muscle stem cell cultures. Scale bar, 100 μm. (D) Diagram illustrating the two selection screens with replicating AAV capsid libraries from (A) that was performed on both pooled primary human skeletal muscle stem cells and human myotubes. (E) Semiquantitative AAV Cap PCR (2.2-Kb product) was performed at each round of each selection screen to demonstrate active replication of AAV library genomes throughout each round of the screen.
    Figure Legend Snippet: Directed Evolution of AAV Capsids by DNA Shuffling and Selection Screening on Primary Human Skeletal Muscle Stem Cells and Myotubes (A) AAV capsid genes from ten parental serotypes (1, 2, 3b, 4, 5, 6, 8, 9_hu14, avian, and bovine) were PCR-amplified, fragmented with DNaseI digestion, and then randomly reassembled through self-priming PCR. Resultant shuffled Cap genes were cloned back into a replication-competent AAV production plasmid via flanking SwaI/NsiI sites downstream of AAV2 Rep . The resultant library production plasmid contained AAV2 ITRs and a modified AAV2 3′ UTR. The AAV library was packaged using standard production protocols as done previously, 41 dot blot titered, and used for selection. (B) Sequential gating scheme Panels [P]1–P4) to isolate CD31 − CD34 − CD45 − ITGB1 + EGFR + primary human skeletal muscle stem cells from surgical specimens. Numbers indicate percentage of total events falling within each gate. (C) Phase contrast and immunofluorescence PAX7 (pink) and DAPI (blue) nuclear staining of purified primary human muscle stem cell cultures. Scale bar, 100 μm. (D) Diagram illustrating the two selection screens with replicating AAV capsid libraries from (A) that was performed on both pooled primary human skeletal muscle stem cells and human myotubes. (E) Semiquantitative AAV Cap PCR (2.2-Kb product) was performed at each round of each selection screen to demonstrate active replication of AAV library genomes throughout each round of the screen.

    Techniques Used: Selection, Polymerase Chain Reaction, Amplification, Clone Assay, Plasmid Preparation, Modification, Dot Blot, Immunofluorescence, Staining, Purification

    10) Product Images from "Efficacy and Mechanisms of Murine Norovirus Inhibition by Pulsed-Light Technology"

    Article Title: Efficacy and Mechanisms of Murine Norovirus Inhibition by Pulsed-Light Technology

    Journal: Applied and Environmental Microbiology

    doi: 10.1128/AEM.03840-14

    Inactivation of MNV-1 on experimentally contaminated polyethylene (A), polyvinyl chloride (B), and stainless steel (C) disks (1.04 cm 2 ) under clean (bars with light cross-hatching) and fouled (bars with dark cross-hatching) conditions after treatment
    Figure Legend Snippet: Inactivation of MNV-1 on experimentally contaminated polyethylene (A), polyvinyl chloride (B), and stainless steel (C) disks (1.04 cm 2 ) under clean (bars with light cross-hatching) and fouled (bars with dark cross-hatching) conditions after treatment

    Techniques Used:

    SDS-PAGE analysis of purified MNV-1 untreated, treated with 2.07 J cm −2 (3 pulses), or treated with 8.98 J cm −2 (13 pulses). Viral proteins were analyzed via 10% SDS-PAGE followed by Coomassie staining. VP1, major capsid protein.
    Figure Legend Snippet: SDS-PAGE analysis of purified MNV-1 untreated, treated with 2.07 J cm −2 (3 pulses), or treated with 8.98 J cm −2 (13 pulses). Viral proteins were analyzed via 10% SDS-PAGE followed by Coomassie staining. VP1, major capsid protein.

    Techniques Used: SDS Page, Purification, Staining

    Activity of pulsed light against MNV-1 in aqueous suspension.
    Figure Legend Snippet: Activity of pulsed light against MNV-1 in aqueous suspension.

    Techniques Used: Activity Assay

    Viral genomic RNA degraded by pulsed light. Viral genomic RNA extracted from treated or untreated purified MNV-1 was loaded onto a 6000 RNA Pico chip and analyzed on a Bioanalyzer with a UV detector. Results are presented in electropherogram form. Light
    Figure Legend Snippet: Viral genomic RNA degraded by pulsed light. Viral genomic RNA extracted from treated or untreated purified MNV-1 was loaded onto a 6000 RNA Pico chip and analyzed on a Bioanalyzer with a UV detector. Results are presented in electropherogram form. Light

    Techniques Used: Purification, Chromatin Immunoprecipitation

    Viral particles disrupted by pulsed light. Infectivity of purified MNV-1 dropped by 5.15 ± 0.18 log 10 after treatment with 2.07 J cm −2 (3 pulses) and was completely eliminated after treatment with 8.98 J cm −2 (13 pulses). Treated
    Figure Legend Snippet: Viral particles disrupted by pulsed light. Infectivity of purified MNV-1 dropped by 5.15 ± 0.18 log 10 after treatment with 2.07 J cm −2 (3 pulses) and was completely eliminated after treatment with 8.98 J cm −2 (13 pulses). Treated

    Techniques Used: Infection, Purification

    Inactivation of MNV-1 in experimentally contaminated hard waters (A) and turbid waters (B) after pulsed-light treatments at 80 mm from a xenon lamp. White bars, 0.69 J cm − ; white bars with black dots, 2.07 J cm −2 ; checkered bars, 3.45
    Figure Legend Snippet: Inactivation of MNV-1 in experimentally contaminated hard waters (A) and turbid waters (B) after pulsed-light treatments at 80 mm from a xenon lamp. White bars, 0.69 J cm − ; white bars with black dots, 2.07 J cm −2 ; checkered bars, 3.45

    Techniques Used:

    11) Product Images from "An adenovirus-derived protein: A novel candidate for anti-diabetic drug development"

    Article Title: An adenovirus-derived protein: A novel candidate for anti-diabetic drug development

    Journal: Biochimie

    doi: 10.1016/j.biochi.2015.12.002

    E4orf1 transiently but reproducibly enhances glucose excursion: C57BL/6J (9 week old) mice on 60% fat diet since 6 week of age were inoculated with pBabe-puro retrovirus (control; black solid lines, n = 6), or with pBabe expressing E4orf1 (black dotted
    Figure Legend Snippet: E4orf1 transiently but reproducibly enhances glucose excursion: C57BL/6J (9 week old) mice on 60% fat diet since 6 week of age were inoculated with pBabe-puro retrovirus (control; black solid lines, n = 6), or with pBabe expressing E4orf1 (black dotted

    Techniques Used: Mouse Assay, Expressing

    Longer duration of high fat diet delays the improvement in GTT induced by E4orf1: Fourteen week old C57BL/6J male mice on 60% fat diet (since 6wk) were inoculated with pBabe-puro (control; black solid lines/bars, n = 6), or with different doses of pBabe
    Figure Legend Snippet: Longer duration of high fat diet delays the improvement in GTT induced by E4orf1: Fourteen week old C57BL/6J male mice on 60% fat diet (since 6wk) were inoculated with pBabe-puro (control; black solid lines/bars, n = 6), or with different doses of pBabe

    Techniques Used: Mouse Assay

    12) Product Images from "HIV-1 Tat Interacts with a Kaposi’s Sarcoma-Associated Herpesvirus Reactivation-Upregulated Antiangiogenic Long Noncoding RNA, LINC00313, and Antagonizes Its Function"

    Article Title: HIV-1 Tat Interacts with a Kaposi’s Sarcoma-Associated Herpesvirus Reactivation-Upregulated Antiangiogenic Long Noncoding RNA, LINC00313, and Antagonizes Its Function

    Journal: Journal of Virology

    doi: 10.1128/JVI.01280-19

    Soluble HIV Tat enhances the invasion and viral production of KSHV lytic infection-reactivated SLK cells. (A) Viability of control and iSLK-BAC16 cells treated with Dox (1 μg/ml) with or without of HIV Tat (0.2 μg/ml) at the indicated time points was assessed using MTT. (B and D) Representative images from cell migration (B) and invasion (D) assays in iSLK-BAC16 cells treated as described for panel A. (C and E) Quantification of cell migration (C) and invasion (E) described in the legend for panels B and D, respectively. (F) Supernatants from iSLK-BAC16 cells treated as described in the legend for panel A were collected at 72 and 96 h, and the viral titers were determined by analyzing the virion-associated DNA levels using TaqMan qPCR. Data shown are means ± standard deviations (SD) ( n = 3). **, P
    Figure Legend Snippet: Soluble HIV Tat enhances the invasion and viral production of KSHV lytic infection-reactivated SLK cells. (A) Viability of control and iSLK-BAC16 cells treated with Dox (1 μg/ml) with or without of HIV Tat (0.2 μg/ml) at the indicated time points was assessed using MTT. (B and D) Representative images from cell migration (B) and invasion (D) assays in iSLK-BAC16 cells treated as described for panel A. (C and E) Quantification of cell migration (C) and invasion (E) described in the legend for panels B and D, respectively. (F) Supernatants from iSLK-BAC16 cells treated as described in the legend for panel A were collected at 72 and 96 h, and the viral titers were determined by analyzing the virion-associated DNA levels using TaqMan qPCR. Data shown are means ± standard deviations (SD) ( n = 3). **, P

    Techniques Used: Infection, MTT Assay, Migration, Real-time Polymerase Chain Reaction

    13) Product Images from "Selection and evaluation of an efficient method for the recovery of viral nucleic acids from complex biologicals"

    Article Title: Selection and evaluation of an efficient method for the recovery of viral nucleic acids from complex biologicals

    Journal: NPJ Vaccines

    doi: 10.1038/s41541-018-0067-3

    Efficiency of separate extraction of DNA and RNA when compared to total nucleic acid extraction using QIAamp® MinElute® Virus Spin kit. DNA and RNA were extracted separately and compared to the total nucleic acid extracted using the QIAamp® MinElute® Virus Spin kit, using qPCR. RNA extraction using the Qiagen RNeasy® Mini kit did not enhance the detection of the two single-stranded RNA viruses (FeLV and RSV), whereas the RNase A digestion added to the Wako DNA Extractor® kit greatly enriched for double-stranded nucleic acid (EBV and Reo3)
    Figure Legend Snippet: Efficiency of separate extraction of DNA and RNA when compared to total nucleic acid extraction using QIAamp® MinElute® Virus Spin kit. DNA and RNA were extracted separately and compared to the total nucleic acid extracted using the QIAamp® MinElute® Virus Spin kit, using qPCR. RNA extraction using the Qiagen RNeasy® Mini kit did not enhance the detection of the two single-stranded RNA viruses (FeLV and RSV), whereas the RNase A digestion added to the Wako DNA Extractor® kit greatly enriched for double-stranded nucleic acid (EBV and Reo3)

    Techniques Used: Real-time Polymerase Chain Reaction, RNA Extraction

    14) Product Images from "A Modified Extraction Method of Circulating Free DNA for Epidermal Growth Factor Receptor Mutation Analysis"

    Article Title: A Modified Extraction Method of Circulating Free DNA for Epidermal Growth Factor Receptor Mutation Analysis

    Journal: Yonsei Medical Journal

    doi: 10.3349/ymj.2012.53.1.132

    Sequencing maps of known EGFR deletion in exon 19 of plasma cfDNA. cfDNA was extracted by modified phenol-chloroform (MPC) method (A) and QIAamp MinElute Virus Spin kit (Qiagen kit) (B). In (A), the height of mutant peak is similar to that of wild-type peak, while the height of mutant peak in (B) is much lower than that of wild-type peak. EGFR, epidermal growth factor receptor.
    Figure Legend Snippet: Sequencing maps of known EGFR deletion in exon 19 of plasma cfDNA. cfDNA was extracted by modified phenol-chloroform (MPC) method (A) and QIAamp MinElute Virus Spin kit (Qiagen kit) (B). In (A), the height of mutant peak is similar to that of wild-type peak, while the height of mutant peak in (B) is much lower than that of wild-type peak. EGFR, epidermal growth factor receptor.

    Techniques Used: Sequencing, Modification, Mutagenesis

    Comparison of fragment distribution of plasma cfDNA extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit).
    Figure Legend Snippet: Comparison of fragment distribution of plasma cfDNA extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit).

    Techniques Used: Modification

    Comparison of cfDNA yield extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit).
    Figure Legend Snippet: Comparison of cfDNA yield extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit).

    Techniques Used: Modification

    Amplification plot of cfDNA extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit) in DxS EGFR mutation test kit. EGFR, epidermal growth factor receptor.
    Figure Legend Snippet: Amplification plot of cfDNA extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit) in DxS EGFR mutation test kit. EGFR, epidermal growth factor receptor.

    Techniques Used: Amplification, Modification, Mutagenesis

    15) Product Images from "A Modified Extraction Method of Circulating Free DNA for Epidermal Growth Factor Receptor Mutation Analysis"

    Article Title: A Modified Extraction Method of Circulating Free DNA for Epidermal Growth Factor Receptor Mutation Analysis

    Journal: Yonsei Medical Journal

    doi: 10.3349/ymj.2012.53.1.132

    Sequencing maps of known EGFR deletion in exon 19 of plasma cfDNA. cfDNA was extracted by modified phenol-chloroform (MPC) method (A) and QIAamp MinElute Virus Spin kit (Qiagen kit) (B). In (A), the height of mutant peak is similar to that of wild-type peak, while the height of mutant peak in (B) is much lower than that of wild-type peak. EGFR, epidermal growth factor receptor.
    Figure Legend Snippet: Sequencing maps of known EGFR deletion in exon 19 of plasma cfDNA. cfDNA was extracted by modified phenol-chloroform (MPC) method (A) and QIAamp MinElute Virus Spin kit (Qiagen kit) (B). In (A), the height of mutant peak is similar to that of wild-type peak, while the height of mutant peak in (B) is much lower than that of wild-type peak. EGFR, epidermal growth factor receptor.

    Techniques Used: Sequencing, Modification, Mutagenesis

    Comparison of fragment distribution of plasma cfDNA extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit).
    Figure Legend Snippet: Comparison of fragment distribution of plasma cfDNA extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit).

    Techniques Used: Modification

    Comparison of cfDNA yield extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit).
    Figure Legend Snippet: Comparison of cfDNA yield extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit).

    Techniques Used: Modification

    Amplification plot of cfDNA extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit) in DxS EGFR mutation test kit. EGFR, epidermal growth factor receptor.
    Figure Legend Snippet: Amplification plot of cfDNA extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit) in DxS EGFR mutation test kit. EGFR, epidermal growth factor receptor.

    Techniques Used: Amplification, Modification, Mutagenesis

    16) Product Images from "Synergistic combination of valproic acid and oncolytic parvovirus H-1PV as a potential therapy against cervical and pancreatic carcinomas"

    Article Title: Synergistic combination of valproic acid and oncolytic parvovirus H-1PV as a potential therapy against cervical and pancreatic carcinomas

    Journal: EMBO Molecular Medicine

    doi: 10.1002/emmm.201302796

    H-1PV/VPA co-treatment leads to complete remission of established HeLa tumours Source data is available for this figure in the Supporting Information. Analysis of tumour growth. Animals ( n = 8 per group) were treated with either PBS (control), VPA or a combination of both agents as described in Materials and Methods Section. Other animal groups are presented in Supporting Information Fig S7 . The data shown represent average tumour volumes with standard deviation bars. Graphical summary of Kaplan–Meier survival analysis of tumour-bearing nude rats treated as indicated. Animals were sacrificed when the tumour reached the maximum tolerable size of 4000 mm 3 according to law. The difference between H-1PV/VPA co-treatment and treatment with H-1PV alone was statistically significant ( p = 0.0002 as calculated with the two-sided log-rank test and adjusted for multiple testing with the Bonferroni method). Other results from the statistical analysis are shown in Supporting Information Table S2. H-1PV/VPA co-treatment induces DNA damage and apoptosis in vivo . Two rats of each treatment group were sacrificed on day 19. Tumours were fixed, sectioned and examined morphologically either by H E staining or by immunochemistry using antibodies against NS1, VP capsid proteins, γ-H2AX (a DNA damage marker), or active cleaved caspase-3 (an apoptosis marker). Scale bar = 50 µm. Higher levels of NS1, VP1 and VP2 viral proteins in tumour samples from VPA co-treated animals. Tumour-bearing animals ( n = 3) treated with H-1PV alone (total virus dose of 3.6 × 10 9 Vg/animal corresponding to 1.5 × 10 8 pfu/animals subdivided into two subdoses given at days 0 and 7) or co-treated with H-1PV and VPA (100 mg/kg) were sacrificed at day 14. Complete tumours were resected, homogenised and lysed. 20 µg of total cellular extracts were analysed by SDS–PAGE for the presence of the viral proteins. Actin was used as a loading control. VPA-enhanced virus multiplication in vivo . Viral particles were recovered from aliquots of tumour extracts from animals treated as in panel D, purified by using the QIAamp MinElute Virus Spin Kit (Qiagen) and quantified by real time qPCR. Columns represent the average amounts of the virus recovered from the tumours of three animals with standard deviation bars. Numbers on top of the columns indicate the fold increase in virus titers in the presence versus absence of VPA.
    Figure Legend Snippet: H-1PV/VPA co-treatment leads to complete remission of established HeLa tumours Source data is available for this figure in the Supporting Information. Analysis of tumour growth. Animals ( n = 8 per group) were treated with either PBS (control), VPA or a combination of both agents as described in Materials and Methods Section. Other animal groups are presented in Supporting Information Fig S7 . The data shown represent average tumour volumes with standard deviation bars. Graphical summary of Kaplan–Meier survival analysis of tumour-bearing nude rats treated as indicated. Animals were sacrificed when the tumour reached the maximum tolerable size of 4000 mm 3 according to law. The difference between H-1PV/VPA co-treatment and treatment with H-1PV alone was statistically significant ( p = 0.0002 as calculated with the two-sided log-rank test and adjusted for multiple testing with the Bonferroni method). Other results from the statistical analysis are shown in Supporting Information Table S2. H-1PV/VPA co-treatment induces DNA damage and apoptosis in vivo . Two rats of each treatment group were sacrificed on day 19. Tumours were fixed, sectioned and examined morphologically either by H E staining or by immunochemistry using antibodies against NS1, VP capsid proteins, γ-H2AX (a DNA damage marker), or active cleaved caspase-3 (an apoptosis marker). Scale bar = 50 µm. Higher levels of NS1, VP1 and VP2 viral proteins in tumour samples from VPA co-treated animals. Tumour-bearing animals ( n = 3) treated with H-1PV alone (total virus dose of 3.6 × 10 9 Vg/animal corresponding to 1.5 × 10 8 pfu/animals subdivided into two subdoses given at days 0 and 7) or co-treated with H-1PV and VPA (100 mg/kg) were sacrificed at day 14. Complete tumours were resected, homogenised and lysed. 20 µg of total cellular extracts were analysed by SDS–PAGE for the presence of the viral proteins. Actin was used as a loading control. VPA-enhanced virus multiplication in vivo . Viral particles were recovered from aliquots of tumour extracts from animals treated as in panel D, purified by using the QIAamp MinElute Virus Spin Kit (Qiagen) and quantified by real time qPCR. Columns represent the average amounts of the virus recovered from the tumours of three animals with standard deviation bars. Numbers on top of the columns indicate the fold increase in virus titers in the presence versus absence of VPA.

    Techniques Used: Standard Deviation, In Vivo, Staining, Marker, SDS Page, Purification, Real-time Polymerase Chain Reaction

    17) Product Images from "Evaluation of the Broad-Range PCR-Electrospray Ionization Mass Spectrometry (PCR/ESI-MS) System and Virus Microarrays for Virus Detection"

    Article Title: Evaluation of the Broad-Range PCR-Electrospray Ionization Mass Spectrometry (PCR/ESI-MS) System and Virus Microarrays for Virus Detection

    Journal: Viruses

    doi: 10.3390/v6051876

    Determination of retrovirus LOD using RT-PCR assays. Total RNA was extracted from each virus dilution to create XMRV and SFV-1 RNA panels (10 0 –10 −7 ) in the absence and presence of 10 5 or 10 4 cell equivalents of Sf9 total nucleic acids as described in Materials and Methods. A subset of the RNA panels (10 −3 –10 −6 ) was subjected to nested RT-PCR assays. An RT minus (−RT) control of the samples shows the absence of residual cellular DNA and the negative, no template control for the PCR shows the absence of contamination in the assay. ( a ) XMRV gag primers; ( b ) SFV-1 gag primers. The size of fragments in the 100 bp marker (M) is indicated in base pairs (bp).
    Figure Legend Snippet: Determination of retrovirus LOD using RT-PCR assays. Total RNA was extracted from each virus dilution to create XMRV and SFV-1 RNA panels (10 0 –10 −7 ) in the absence and presence of 10 5 or 10 4 cell equivalents of Sf9 total nucleic acids as described in Materials and Methods. A subset of the RNA panels (10 −3 –10 −6 ) was subjected to nested RT-PCR assays. An RT minus (−RT) control of the samples shows the absence of residual cellular DNA and the negative, no template control for the PCR shows the absence of contamination in the assay. ( a ) XMRV gag primers; ( b ) SFV-1 gag primers. The size of fragments in the 100 bp marker (M) is indicated in base pairs (bp).

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Marker

    18) Product Images from "An adenovirus-derived protein: A novel candidate for anti-diabetic drug development"

    Article Title: An adenovirus-derived protein: A novel candidate for anti-diabetic drug development

    Journal: Biochimie

    doi: 10.1016/j.biochi.2015.12.002

    E4orf1 transiently but reproducibly enhances glucose excursion: C57BL/6J (9 week old) mice on 60% fat diet since 6 week of age were inoculated with pBabe-puro retrovirus (control; black solid lines, n = 6), or with pBabe expressing E4orf1 (black dotted
    Figure Legend Snippet: E4orf1 transiently but reproducibly enhances glucose excursion: C57BL/6J (9 week old) mice on 60% fat diet since 6 week of age were inoculated with pBabe-puro retrovirus (control; black solid lines, n = 6), or with pBabe expressing E4orf1 (black dotted

    Techniques Used: Mouse Assay, Expressing

    A booster dose of E4orf1 does not extend the duration of enhancement in glucose clearance: Nine week old C57BL/6J (9 wk old) male mice on 60% HF diet since 6 wk of age were inoculated with control retrovirus (black solid lines, n = 6), or with E4orf1
    Figure Legend Snippet: A booster dose of E4orf1 does not extend the duration of enhancement in glucose clearance: Nine week old C57BL/6J (9 wk old) male mice on 60% HF diet since 6 wk of age were inoculated with control retrovirus (black solid lines, n = 6), or with E4orf1

    Techniques Used: Mouse Assay

    E4orf1 improves glucose clearance without increasing insulin sensitivity, production or secretion: Insulin tolerance test (ITT) was performed 1 and 2 week post initial inoculation of control retrovirus (black solid lines, n = 3), or with E4orf1 (black
    Figure Legend Snippet: E4orf1 improves glucose clearance without increasing insulin sensitivity, production or secretion: Insulin tolerance test (ITT) was performed 1 and 2 week post initial inoculation of control retrovirus (black solid lines, n = 3), or with E4orf1 (black

    Techniques Used:

    19) Product Images from "Fast-Seq: A Simple Method for Rapid and Inexpensive Validation of Packaged Single-Stranded Adeno-Associated Viral Genomes in Academic Settings"

    Article Title: Fast-Seq: A Simple Method for Rapid and Inexpensive Validation of Packaged Single-Stranded Adeno-Associated Viral Genomes in Academic Settings

    Journal: Human Gene Therapy Methods

    doi: 10.1089/hgtb.2019.110

    Overview of Fast-Seq, a Tn5-based packaged ssAAV genome sequencing method. Preparations of rAAV undergo ssDNA extraction, second-strand synthesis to generate dsDNA, tagmentation with adapter-loaded Tn5 transposomes, indexing, QC validation, short-read sequencing on Illumina MiSeq, mapping reads to the transfer vector plasmid used to generate the input rAAV, and sequence validation of the packaged genome. An example packaged genome mutation (*) is shown at position 7. dsAAV, double-stranded AAV; dsDNA, double-stranded DNA; rAAV, recombinant AAV; ssAAV, single-stranded adeno-associated virus; Tn5, transposase.
    Figure Legend Snippet: Overview of Fast-Seq, a Tn5-based packaged ssAAV genome sequencing method. Preparations of rAAV undergo ssDNA extraction, second-strand synthesis to generate dsDNA, tagmentation with adapter-loaded Tn5 transposomes, indexing, QC validation, short-read sequencing on Illumina MiSeq, mapping reads to the transfer vector plasmid used to generate the input rAAV, and sequence validation of the packaged genome. An example packaged genome mutation (*) is shown at position 7. dsAAV, double-stranded AAV; dsDNA, double-stranded DNA; rAAV, recombinant AAV; ssAAV, single-stranded adeno-associated virus; Tn5, transposase.

    Techniques Used: Sequencing, Plasmid Preparation, Mutagenesis, Recombinant

    20) Product Images from "A Modified Extraction Method of Circulating Free DNA for Epidermal Growth Factor Receptor Mutation Analysis"

    Article Title: A Modified Extraction Method of Circulating Free DNA for Epidermal Growth Factor Receptor Mutation Analysis

    Journal: Yonsei Medical Journal

    doi: 10.3349/ymj.2012.53.1.132

    Sequencing maps of known EGFR deletion in exon 19 of plasma cfDNA. cfDNA was extracted by modified phenol-chloroform (MPC) method (A) and QIAamp MinElute Virus Spin kit (Qiagen kit) (B). In (A), the height of mutant peak is similar to that of wild-type peak, while the height of mutant peak in (B) is much lower than that of wild-type peak. EGFR, epidermal growth factor receptor.
    Figure Legend Snippet: Sequencing maps of known EGFR deletion in exon 19 of plasma cfDNA. cfDNA was extracted by modified phenol-chloroform (MPC) method (A) and QIAamp MinElute Virus Spin kit (Qiagen kit) (B). In (A), the height of mutant peak is similar to that of wild-type peak, while the height of mutant peak in (B) is much lower than that of wild-type peak. EGFR, epidermal growth factor receptor.

    Techniques Used: Sequencing, Modification, Mutagenesis

    Comparison of fragment distribution of plasma cfDNA extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit).
    Figure Legend Snippet: Comparison of fragment distribution of plasma cfDNA extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit).

    Techniques Used: Modification

    Comparison of cfDNA yield extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit).
    Figure Legend Snippet: Comparison of cfDNA yield extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit).

    Techniques Used: Modification

    Amplification plot of cfDNA extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit) in DxS EGFR mutation test kit. EGFR, epidermal growth factor receptor.
    Figure Legend Snippet: Amplification plot of cfDNA extracted by modified phenol-chloroform (MPC) method and QIAamp MinElute Virus Spin kit (Qiagen kit) in DxS EGFR mutation test kit. EGFR, epidermal growth factor receptor.

    Techniques Used: Amplification, Modification, Mutagenesis

    Related Articles

    Modification:

    Article Title: A Modified Extraction Method of Circulating Free DNA for Epidermal Growth Factor Receptor Mutation Analysis
    Article Snippet: .. The yield of plasma cfDNA extracted by our modified phenol-chloroform (MPC) method from non-small-cell lung cancer (NSCLC) patients was compared with that by QIAamp MinElute Virus Spin kit (Qiagen kit) as control, using the Wilcoxon rank-sum test. .. TaqMan quantitative polymerase chain reaction (qPCR) assays were used to quantify the plasma cfDNA extracted.

    Real-time Polymerase Chain Reaction:

    Article Title: Selection and evaluation of an efficient method for the recovery of viral nucleic acids from complex biologicals
    Article Snippet: .. Methods that were identified as candidates for efficient viral nucleic acid recovery for different types of viral nucleic acids, such as the Invitrogen PureLink™ Virus RNA/DNA kit and the QIAGEN QIAamp® MinElute® Virus Spin kit, were assessed for repeatability (n = 6) and were found to be within the same order of magnitude, between extractions from the same kit, for total copy number of viral nucleic acid when controlled for the total mass (111 ng) of extracted nucleic acid used for first-strand cDNA synthesis followed by qPCR. ..

    Article Title: Selection and evaluation of an efficient method for the recovery of viral nucleic acids from complex biologicals
    Article Snippet: .. The efficiency of nucleic acid extraction was measured by qPCR and the data are presented as fold change in comparison to the QIAGEN QIAamp® MinElute® Virus Spin kit for total nucleic acid extraction. .. Compared to total nucleic acid extraction using the QIAamp® MinElute® Virus Spin kit, RNA extraction using DNase I did not enhance the recovery of the two single-stranded RNA viruses, however, the RNase A digestion step enriched DNA extraction for double-stranded nucleic acids (both DNA and RNA; Fig. ).

    Mutagenesis:

    Article Title: A Modified Extraction Method of Circulating Free DNA for Epidermal Growth Factor Receptor Mutation Analysis
    Article Snippet: .. Since QIAamp MinElute Virus Spin kit, in comparison with several other commercially available kits, has been reported to yield the highest amount of cfDNA, we compared our MPC method to Qiagen kit method on cfDNA extraction yield and the impact of different extraction method on EGFR mutation analysis. .. Subjects and samples A total of 25 patients with advanced lung adenocarcinoma were recruited from No.3 People's Hospital, School of Medicine, Shanghai Jiaotong University, China between March and September 2009.

    Purification:

    Article Title: Synergistic combination of valproic acid and oncolytic parvovirus H-1PV as a potential therapy against cervical and pancreatic carcinomas
    Article Snippet: .. Viral genomic DNA was purified from cell lysates by using the QiaAmp MinElute virus kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions. .. Quantification of viral DNA was carried out by real-time qPCR, as previously described (El-Andaloussi et al, ).

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    Qiagen qiaamp minelute virus spin kit
    Efficiency of separate extraction of DNA and RNA when compared to total nucleic acid extraction using <t>QIAamp®</t> <t>MinElute®</t> Virus Spin kit. DNA and RNA were extracted separately and compared to the total nucleic acid extracted using the QIAamp® MinElute® Virus Spin kit, using qPCR. RNA extraction using the Qiagen RNeasy® Mini kit did not enhance the detection of the two single-stranded RNA viruses (FeLV and RSV), whereas the RNase A digestion added to the Wako DNA Extractor® kit greatly enriched for double-stranded nucleic acid (EBV and Reo3)
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    Efficiency of separate extraction of DNA and RNA when compared to total nucleic acid extraction using QIAamp® MinElute® Virus Spin kit. DNA and RNA were extracted separately and compared to the total nucleic acid extracted using the QIAamp® MinElute® Virus Spin kit, using qPCR. RNA extraction using the Qiagen RNeasy® Mini kit did not enhance the detection of the two single-stranded RNA viruses (FeLV and RSV), whereas the RNase A digestion added to the Wako DNA Extractor® kit greatly enriched for double-stranded nucleic acid (EBV and Reo3)

    Journal: NPJ Vaccines

    Article Title: Selection and evaluation of an efficient method for the recovery of viral nucleic acids from complex biologicals

    doi: 10.1038/s41541-018-0067-3

    Figure Lengend Snippet: Efficiency of separate extraction of DNA and RNA when compared to total nucleic acid extraction using QIAamp® MinElute® Virus Spin kit. DNA and RNA were extracted separately and compared to the total nucleic acid extracted using the QIAamp® MinElute® Virus Spin kit, using qPCR. RNA extraction using the Qiagen RNeasy® Mini kit did not enhance the detection of the two single-stranded RNA viruses (FeLV and RSV), whereas the RNase A digestion added to the Wako DNA Extractor® kit greatly enriched for double-stranded nucleic acid (EBV and Reo3)

    Article Snippet: The data are presented as fold change in comparison to the QIAGEN QIAamp® MinElute® Virus Spin kit.

    Techniques: Real-time Polymerase Chain Reaction, RNA Extraction