dna input  (New England Biolabs)


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  • 99
    Name:
    NEBNext Ultra Ligation Module
    Description:
    NEBNext Ultra Ligation Module 96 rxns
    Catalog Number:
    e7445l
    Price:
    1195
    Size:
    96 rxns
    Category:
    DNA Template Preparation for PCR
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    Structured Review

    New England Biolabs dna input
    NEBNext Ultra Ligation Module
    NEBNext Ultra Ligation Module 96 rxns
    https://www.bioz.com/result/dna input/product/New England Biolabs
    Average 99 stars, based on 8 article reviews
    Price from $9.99 to $1999.99
    dna input - by Bioz Stars, 2020-07
    99/100 stars

    Images

    1) Product Images from "Same-Day Diagnostic and Surveillance Data for Tuberculosis via Whole-Genome Sequencing of Direct Respiratory Samples"

    Article Title: Same-Day Diagnostic and Surveillance Data for Tuberculosis via Whole-Genome Sequencing of Direct Respiratory Samples

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.02483-16

    Timelines. We compared the method of Brown et al. with the results of this study obtained with Illumina MiSeq and MiniSeq and ONT MinION. We assumed that no step of the process can be initiated after 6 p.m. or before 8 a.m. The method of Brown et al. has a rapid extraction step but also a 20-h overnight enrichment step, resulting in a 50-h turnaround time. In our study, we did 27-h MiSeq runs (paired 150-bp reads), but since Mykrobe is k-mer based, a 16-h run (paired 75-bp reads) would give equivalent results; we therefore display that potential timeline here. The DNA extraction process was updated for the MiniSeq and MinION experiments, removing the ethanol precipitation step. In normal use, this would take 3 h. The 1.5-h orange rectangle on the MinION time lines includes both PCR and the 10-min sample preparation step. In this experiment, since we used spiked BCG DNA in sputum, we did not use a human depletion step, thus taking only 2 h. This image is intended to show comparable real-use time lines, and so the MiniSeq/MinION time lines are shown with 3-h extraction steps. MiniSeq enables a 16-h turnaround time by sequencing for only 7 h. R9 MinION also delivers sub-24-h results but requires one flow cell per sample. R9.4 MinION gives a 12.5-h turnaround time (6 h of sequencing with real-time [i.e., simultaneous] base calling when used on a single sample).
    Figure Legend Snippet: Timelines. We compared the method of Brown et al. with the results of this study obtained with Illumina MiSeq and MiniSeq and ONT MinION. We assumed that no step of the process can be initiated after 6 p.m. or before 8 a.m. The method of Brown et al. has a rapid extraction step but also a 20-h overnight enrichment step, resulting in a 50-h turnaround time. In our study, we did 27-h MiSeq runs (paired 150-bp reads), but since Mykrobe is k-mer based, a 16-h run (paired 75-bp reads) would give equivalent results; we therefore display that potential timeline here. The DNA extraction process was updated for the MiniSeq and MinION experiments, removing the ethanol precipitation step. In normal use, this would take 3 h. The 1.5-h orange rectangle on the MinION time lines includes both PCR and the 10-min sample preparation step. In this experiment, since we used spiked BCG DNA in sputum, we did not use a human depletion step, thus taking only 2 h. This image is intended to show comparable real-use time lines, and so the MiniSeq/MinION time lines are shown with 3-h extraction steps. MiniSeq enables a 16-h turnaround time by sequencing for only 7 h. R9 MinION also delivers sub-24-h results but requires one flow cell per sample. R9.4 MinION gives a 12.5-h turnaround time (6 h of sequencing with real-time [i.e., simultaneous] base calling when used on a single sample).

    Techniques Used: DNA Extraction, Ethanol Precipitation, Polymerase Chain Reaction, Sample Prep, Sequencing, Flow Cytometry

    2) Product Images from "Quantitation of next generation sequencing library preparation protocol efficiencies using droplet digital PCR assays - a systematic comparison of DNA library preparation kits for Illumina sequencing"

    Article Title: Quantitation of next generation sequencing library preparation protocol efficiencies using droplet digital PCR assays - a systematic comparison of DNA library preparation kits for Illumina sequencing

    Journal: BMC Genomics

    doi: 10.1186/s12864-016-2757-4

    Example of yield measurements obtained with the NEBNext kit when comparing the amount of DNA at each step versus the initial DNA input (500 ng). Blue bars correspond to values measured using the PhiX specific primers and reflect the DNA loss at each step mainly due to pipetting and bead clean up. The orange bar corresponds to the value measured with the adaptor specific primers and reflects the amount of DNA in the sample bearing adaptor after the ligation step in comparison with the initial DNA input. The green bar corresponds to the value measured with the P5/P7 primers and reflects the amount of sequencable DNA in the sample at the end of the library preparation
    Figure Legend Snippet: Example of yield measurements obtained with the NEBNext kit when comparing the amount of DNA at each step versus the initial DNA input (500 ng). Blue bars correspond to values measured using the PhiX specific primers and reflect the DNA loss at each step mainly due to pipetting and bead clean up. The orange bar corresponds to the value measured with the adaptor specific primers and reflects the amount of DNA in the sample bearing adaptor after the ligation step in comparison with the initial DNA input. The green bar corresponds to the value measured with the P5/P7 primers and reflects the amount of sequencable DNA in the sample at the end of the library preparation

    Techniques Used: Ligation

    3) Product Images from "Quantitation of next generation sequencing library preparation protocol efficiencies using droplet digital PCR assays - a systematic comparison of DNA library preparation kits for Illumina sequencing"

    Article Title: Quantitation of next generation sequencing library preparation protocol efficiencies using droplet digital PCR assays - a systematic comparison of DNA library preparation kits for Illumina sequencing

    Journal: BMC Genomics

    doi: 10.1186/s12864-016-2757-4

    Example of yield measurements obtained with the NEBNext kit when comparing the amount of DNA at each step versus the initial DNA input (500 ng). Blue bars correspond to values measured using the PhiX specific primers and reflect the DNA loss at each step mainly due to pipetting and bead clean up. The orange bar corresponds to the value measured with the adaptor specific primers and reflects the amount of DNA in the sample bearing adaptor after the ligation step in comparison with the initial DNA input. The green bar corresponds to the value measured with the P5/P7 primers and reflects the amount of sequencable DNA in the sample at the end of the library preparation
    Figure Legend Snippet: Example of yield measurements obtained with the NEBNext kit when comparing the amount of DNA at each step versus the initial DNA input (500 ng). Blue bars correspond to values measured using the PhiX specific primers and reflect the DNA loss at each step mainly due to pipetting and bead clean up. The orange bar corresponds to the value measured with the adaptor specific primers and reflects the amount of DNA in the sample bearing adaptor after the ligation step in comparison with the initial DNA input. The green bar corresponds to the value measured with the P5/P7 primers and reflects the amount of sequencable DNA in the sample at the end of the library preparation

    Techniques Used: Ligation

    Related Articles

    Sequencing:

    Article Title: Illuminating Choices for Library Prep: A Comparison of Library Preparation Methods for Whole Genome Sequencing of Cryptococcus neoformans Using Illumina HiSeq
    Article Snippet: .. NEBNext Ultra also boasts advantages such as low inputs of gDNA (5 ng), and creates indexed libraries suitable for the Illumina platform sequencing machines, and as such, is marketed as a cheaper alternative to Illumina. .. The human-infecting pathogenic fungus C. neoformans var. grubii (Cng henceforth) is routinely sequenced in our laboratory, with DNA extraction methods optimised for whole-genome sequencing applications.

    Article Title: Facile, High Quality Sequencing of Bacterial Genomes from Small Amounts of DNA
    Article Snippet: .. Modern library preparation methods for next generation sequencing technologies, as represented by NEBNext Ultra, are enabling bacterial genome sequencing from very small input amounts of genomic DNA. ..

    Polymerase Chain Reaction:

    Article Title: Quantitation of next generation sequencing library preparation protocol efficiencies using droplet digital PCR assays - a systematic comparison of DNA library preparation kits for Illumina sequencing
    Article Snippet: .. We present here the quantitative comparison of 9 kits: NEBNext and NEBNext Ultra from New England Biolabs, SureSelectXT from Agilent, Truseq Nano and Truseq DNA PCR-free from Illumina, Accel-NGS 1S and Accel-NGS 2S from Swift Biosciences, and KAPA Hyper and KAPA HyperPlus from KAPA Biosystems. ..

    Next-Generation Sequencing:

    Article Title: Facile, High Quality Sequencing of Bacterial Genomes from Small Amounts of DNA
    Article Snippet: .. Modern library preparation methods for next generation sequencing technologies, as represented by NEBNext Ultra, are enabling bacterial genome sequencing from very small input amounts of genomic DNA. ..

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    New England Biolabs input dna
    Chromatin structures at CTCF sites around the β-globin locus having deletion of GATA-1 binding motifs. ChIP was performed with antibodies specific to CTCF (A), Rad21 (B), histone H3 (C) and <t>H3K27ac</t> (D) in control cells (Con) and HS3ΔΔGA and HS2ΔGA clones. Amounts of immunoprecipitated <t>DNA</t> were determined as described in Fig 1C . The mouse Necdin gene served as an internal control. The results are the means of four independent experiments ± SEM. *P
    Input Dna, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 94/100, based on 114 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/input dna/product/New England Biolabs
    Average 94 stars, based on 114 article reviews
    Price from $9.99 to $1999.99
    input dna - by Bioz Stars, 2020-07
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    90
    New England Biolabs input bacmid dna
    Subcellular localization of total actin and F-actin in Sf9 cells transfected with different recombinant viruses. (A) Total actin localization. Sf9 cells were transfected with vAc54KO, vAc54:2HA, or v38KKO <t>bacmid</t> <t>DNA.</t> At 36 h p.t., the cells were fixed,
    Input Bacmid Dna, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/input bacmid dna/product/New England Biolabs
    Average 90 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    input bacmid dna - by Bioz Stars, 2020-07
    90/100 stars
      Buy from Supplier

    Image Search Results


    Chromatin structures at CTCF sites around the β-globin locus having deletion of GATA-1 binding motifs. ChIP was performed with antibodies specific to CTCF (A), Rad21 (B), histone H3 (C) and H3K27ac (D) in control cells (Con) and HS3ΔΔGA and HS2ΔGA clones. Amounts of immunoprecipitated DNA were determined as described in Fig 1C . The mouse Necdin gene served as an internal control. The results are the means of four independent experiments ± SEM. *P

    Journal: bioRxiv

    Article Title: GATA-1-dependent histone H3K27ac mediates erythroid cell-specific interaction between CTCF sites

    doi: 10.1101/2020.05.14.095547

    Figure Lengend Snippet: Chromatin structures at CTCF sites around the β-globin locus having deletion of GATA-1 binding motifs. ChIP was performed with antibodies specific to CTCF (A), Rad21 (B), histone H3 (C) and H3K27ac (D) in control cells (Con) and HS3ΔΔGA and HS2ΔGA clones. Amounts of immunoprecipitated DNA were determined as described in Fig 1C . The mouse Necdin gene served as an internal control. The results are the means of four independent experiments ± SEM. *P

    Article Snippet: ChIP DNA library preparation and sequencing ChIP DNA and input DNA (10 ng for H3K27ac, 0.5 ng for CTCF and 10 ng for input) were processed using NEBNext ChIP-seq library (New England Biolabs) with manufacturer’s instructions.

    Techniques: Binding Assay, Chromatin Immunoprecipitation, Clone Assay, Immunoprecipitation

    Chromatin interaction between CTCF sites around the β-globin locus in TSA treated MEL/ch11 cells. (A) MEL/ch11 cells were treated with 25 ng/ml of TSA for 6 h or 24 h. Histone H3 acetylated at K27 was detected by Western blotting in nuclear extract from control cells (Con) and cells treated with TSA. Histone H3 was used as an experimental control. (B) H3K27ac was determined in CTCF sites around the β-globin locus by ChIP. DNA immunoprecipitated by H3K27ac antibodies were quantitatively compared with DNA immunoprecipitated by H3, and then normalized to value in control cells. Normal IgG (IgG) was used as an experimental negative control. (C) Relative cross-linking frequencies was determined between CTCF sites around the β-globin locus in 3C assay as described in Fig 1E . Fragments containing C5 and C3 were used as anchors. Occupancies of CTCF (D) and Rad21 (E) were determined at the CTCF sites by ChIP. Results are presented as the means ± SEM of four to six independent experiments in ChIP and 3C assay. *P

    Journal: bioRxiv

    Article Title: GATA-1-dependent histone H3K27ac mediates erythroid cell-specific interaction between CTCF sites

    doi: 10.1101/2020.05.14.095547

    Figure Lengend Snippet: Chromatin interaction between CTCF sites around the β-globin locus in TSA treated MEL/ch11 cells. (A) MEL/ch11 cells were treated with 25 ng/ml of TSA for 6 h or 24 h. Histone H3 acetylated at K27 was detected by Western blotting in nuclear extract from control cells (Con) and cells treated with TSA. Histone H3 was used as an experimental control. (B) H3K27ac was determined in CTCF sites around the β-globin locus by ChIP. DNA immunoprecipitated by H3K27ac antibodies were quantitatively compared with DNA immunoprecipitated by H3, and then normalized to value in control cells. Normal IgG (IgG) was used as an experimental negative control. (C) Relative cross-linking frequencies was determined between CTCF sites around the β-globin locus in 3C assay as described in Fig 1E . Fragments containing C5 and C3 were used as anchors. Occupancies of CTCF (D) and Rad21 (E) were determined at the CTCF sites by ChIP. Results are presented as the means ± SEM of four to six independent experiments in ChIP and 3C assay. *P

    Article Snippet: ChIP DNA library preparation and sequencing ChIP DNA and input DNA (10 ng for H3K27ac, 0.5 ng for CTCF and 10 ng for input) were processed using NEBNext ChIP-seq library (New England Biolabs) with manufacturer’s instructions.

    Techniques: Western Blot, Chromatin Immunoprecipitation, Immunoprecipitation, Negative Control

    Timelines. We compared the method of Brown et al. with the results of this study obtained with Illumina MiSeq and MiniSeq and ONT MinION. We assumed that no step of the process can be initiated after 6 p.m. or before 8 a.m. The method of Brown et al. has a rapid extraction step but also a 20-h overnight enrichment step, resulting in a 50-h turnaround time. In our study, we did 27-h MiSeq runs (paired 150-bp reads), but since Mykrobe is k-mer based, a 16-h run (paired 75-bp reads) would give equivalent results; we therefore display that potential timeline here. The DNA extraction process was updated for the MiniSeq and MinION experiments, removing the ethanol precipitation step. In normal use, this would take 3 h. The 1.5-h orange rectangle on the MinION time lines includes both PCR and the 10-min sample preparation step. In this experiment, since we used spiked BCG DNA in sputum, we did not use a human depletion step, thus taking only 2 h. This image is intended to show comparable real-use time lines, and so the MiniSeq/MinION time lines are shown with 3-h extraction steps. MiniSeq enables a 16-h turnaround time by sequencing for only 7 h. R9 MinION also delivers sub-24-h results but requires one flow cell per sample. R9.4 MinION gives a 12.5-h turnaround time (6 h of sequencing with real-time [i.e., simultaneous] base calling when used on a single sample).

    Journal: Journal of Clinical Microbiology

    Article Title: Same-Day Diagnostic and Surveillance Data for Tuberculosis via Whole-Genome Sequencing of Direct Respiratory Samples

    doi: 10.1128/JCM.02483-16

    Figure Lengend Snippet: Timelines. We compared the method of Brown et al. with the results of this study obtained with Illumina MiSeq and MiniSeq and ONT MinION. We assumed that no step of the process can be initiated after 6 p.m. or before 8 a.m. The method of Brown et al. has a rapid extraction step but also a 20-h overnight enrichment step, resulting in a 50-h turnaround time. In our study, we did 27-h MiSeq runs (paired 150-bp reads), but since Mykrobe is k-mer based, a 16-h run (paired 75-bp reads) would give equivalent results; we therefore display that potential timeline here. The DNA extraction process was updated for the MiniSeq and MinION experiments, removing the ethanol precipitation step. In normal use, this would take 3 h. The 1.5-h orange rectangle on the MinION time lines includes both PCR and the 10-min sample preparation step. In this experiment, since we used spiked BCG DNA in sputum, we did not use a human depletion step, thus taking only 2 h. This image is intended to show comparable real-use time lines, and so the MiniSeq/MinION time lines are shown with 3-h extraction steps. MiniSeq enables a 16-h turnaround time by sequencing for only 7 h. R9 MinION also delivers sub-24-h results but requires one flow cell per sample. R9.4 MinION gives a 12.5-h turnaround time (6 h of sequencing with real-time [i.e., simultaneous] base calling when used on a single sample).

    Article Snippet: These samples, along with pure BCG DNA, were prepared in accordance with ONTs PCR-based protocol for low-input libraries (DP006_revB_14Aug2015), with modified primers supplied by ONT, a 20-ng DNA input into the PCR, and LongAmp Taq 2× master mix (New England BioLabs, USA).

    Techniques: DNA Extraction, Ethanol Precipitation, Polymerase Chain Reaction, Sample Prep, Sequencing, Flow Cytometry

    Attenuating nuclease activity of SpCas9. ( a ) Four PAM-interacting amino acids neighboring the nGG PAM (magenta) in the structure of SpCas9 7 . Arginines at positions 1333 and 1335 were mutated to attenuate DNA-binding affinity of SpCas9. ( b ) Activity profile of SpCas9 (blue) or SpCas9-Zif268 (red) bearing lysine or serine substitutions at positions 1333 or 1335 in the PAM interaction domain in comparison to wild-type (WT) SpCas9. Reporter assays were performed in HEK293T cells. Bar heights represent means from three independent biological replicates performed on different days. Error bars indicate standard error of the mean. ( c ) T7 Endonuclease I (T7EI) assays on PCR products spanning a genomic target site (underlined) with an NGG PAM (magenta) and neighboring Zif268 site (orange) for SpCas9 or SpCas9 mutants with or without a Zif268 fusion. For SpCas9 MT2 SpCas9 MT3 , robust nuclease activity is only observed when Zif268 is fused to the C-terminus. The gel image is representative of T7EI assays at this genomic target site, where cleaved products are noted by magenta arrowheads. ( d ) Quantification of average T7EI-based lesion rates at the PLXNB2 locus from three independent biological replicates performed on different days in HEK293T cells ( Supplementary Fig. 7 ). Error bars indicate standard error of the mean.

    Journal: Nature methods

    Article Title: DNA-binding domain fusions enhance the targeting range and precision of Cas9

    doi: 10.1038/nmeth.3624

    Figure Lengend Snippet: Attenuating nuclease activity of SpCas9. ( a ) Four PAM-interacting amino acids neighboring the nGG PAM (magenta) in the structure of SpCas9 7 . Arginines at positions 1333 and 1335 were mutated to attenuate DNA-binding affinity of SpCas9. ( b ) Activity profile of SpCas9 (blue) or SpCas9-Zif268 (red) bearing lysine or serine substitutions at positions 1333 or 1335 in the PAM interaction domain in comparison to wild-type (WT) SpCas9. Reporter assays were performed in HEK293T cells. Bar heights represent means from three independent biological replicates performed on different days. Error bars indicate standard error of the mean. ( c ) T7 Endonuclease I (T7EI) assays on PCR products spanning a genomic target site (underlined) with an NGG PAM (magenta) and neighboring Zif268 site (orange) for SpCas9 or SpCas9 mutants with or without a Zif268 fusion. For SpCas9 MT2 SpCas9 MT3 , robust nuclease activity is only observed when Zif268 is fused to the C-terminus. The gel image is representative of T7EI assays at this genomic target site, where cleaved products are noted by magenta arrowheads. ( d ) Quantification of average T7EI-based lesion rates at the PLXNB2 locus from three independent biological replicates performed on different days in HEK293T cells ( Supplementary Fig. 7 ). Error bars indicate standard error of the mean.

    Article Snippet: 50ng input DNA is PCR-amplified using “T7EI primers” that are specific for each genomic region ( ) with Phusion High Fidelity DNA Polymerase (New England Biolabs): (98°C, 15s; 67°C, 25s; 72°C, 18s) for 30 cycles.

    Techniques: Activity Assay, Binding Assay, Polymerase Chain Reaction

    Subcellular localization of total actin and F-actin in Sf9 cells transfected with different recombinant viruses. (A) Total actin localization. Sf9 cells were transfected with vAc54KO, vAc54:2HA, or v38KKO bacmid DNA. At 36 h p.t., the cells were fixed,

    Journal: Journal of Virology

    Article Title: The Autographa californica Multiple Nucleopolyhedrovirus ac54 Gene Is Crucial for Localization of the Major Capsid Protein VP39 at the Site of Nucleocapsid Assembly

    doi: 10.1128/JVI.02885-15

    Figure Lengend Snippet: Subcellular localization of total actin and F-actin in Sf9 cells transfected with different recombinant viruses. (A) Total actin localization. Sf9 cells were transfected with vAc54KO, vAc54:2HA, or v38KKO bacmid DNA. At 36 h p.t., the cells were fixed,

    Article Snippet: To eliminate input bacmid DNA, 1 μg of DNA was digested with 20 units of the DpnI restriction enzyme (NEB) overnight in a 40-μl reaction volume. qPCR was performed using Hot Start PCR master mix III (Chaoshi-Bio) and primers targeting a 100-bp region of the gp41 gene under conditions described previously ( ).

    Techniques: Transfection, Recombinant

    Subcellular localization of the minor capsid proteins PP78/83, BV/ODV-C42 (C42), and 38K, as indicated. Sf9 cells were transfected with vAc54KO, vAc54:2HA, or v38KKO bacmid DNA and fixed at 36 h p.t. (A) Transfected cells were incubated with antisera

    Journal: Journal of Virology

    Article Title: The Autographa californica Multiple Nucleopolyhedrovirus ac54 Gene Is Crucial for Localization of the Major Capsid Protein VP39 at the Site of Nucleocapsid Assembly

    doi: 10.1128/JVI.02885-15

    Figure Lengend Snippet: Subcellular localization of the minor capsid proteins PP78/83, BV/ODV-C42 (C42), and 38K, as indicated. Sf9 cells were transfected with vAc54KO, vAc54:2HA, or v38KKO bacmid DNA and fixed at 36 h p.t. (A) Transfected cells were incubated with antisera

    Article Snippet: To eliminate input bacmid DNA, 1 μg of DNA was digested with 20 units of the DpnI restriction enzyme (NEB) overnight in a 40-μl reaction volume. qPCR was performed using Hot Start PCR master mix III (Chaoshi-Bio) and primers targeting a 100-bp region of the gp41 gene under conditions described previously ( ).

    Techniques: Transfection, Incubation

    Subcellular localization of the major capsid protein VP39. Sf9 cells were transfected with vAc54KO (A), vAc54:2HA (B), or v38KKO (C) bacmid DNA. Transfected cells were fixed at 36 h p.t. and incubated with antisera against VP39 (rabbit) and IE-1 (mouse)

    Journal: Journal of Virology

    Article Title: The Autographa californica Multiple Nucleopolyhedrovirus ac54 Gene Is Crucial for Localization of the Major Capsid Protein VP39 at the Site of Nucleocapsid Assembly

    doi: 10.1128/JVI.02885-15

    Figure Lengend Snippet: Subcellular localization of the major capsid protein VP39. Sf9 cells were transfected with vAc54KO (A), vAc54:2HA (B), or v38KKO (C) bacmid DNA. Transfected cells were fixed at 36 h p.t. and incubated with antisera against VP39 (rabbit) and IE-1 (mouse)

    Article Snippet: To eliminate input bacmid DNA, 1 μg of DNA was digested with 20 units of the DpnI restriction enzyme (NEB) overnight in a 40-μl reaction volume. qPCR was performed using Hot Start PCR master mix III (Chaoshi-Bio) and primers targeting a 100-bp region of the gp41 gene under conditions described previously ( ).

    Techniques: Transfection, Incubation

    Characterization of the ac54 -knockout bacmid. (A) Strategy for the construction of the recombinant viruses vAc54KO, vAc54:2HA, and vAcWT. (B) Sf9 cells were transfected with vAc54KO, vAc54:2HA, and vAcWT bacmid DNA and observed at 72 h p.t. (C) Virus

    Journal: Journal of Virology

    Article Title: The Autographa californica Multiple Nucleopolyhedrovirus ac54 Gene Is Crucial for Localization of the Major Capsid Protein VP39 at the Site of Nucleocapsid Assembly

    doi: 10.1128/JVI.02885-15

    Figure Lengend Snippet: Characterization of the ac54 -knockout bacmid. (A) Strategy for the construction of the recombinant viruses vAc54KO, vAc54:2HA, and vAcWT. (B) Sf9 cells were transfected with vAc54KO, vAc54:2HA, and vAcWT bacmid DNA and observed at 72 h p.t. (C) Virus

    Article Snippet: To eliminate input bacmid DNA, 1 μg of DNA was digested with 20 units of the DpnI restriction enzyme (NEB) overnight in a 40-μl reaction volume. qPCR was performed using Hot Start PCR master mix III (Chaoshi-Bio) and primers targeting a 100-bp region of the gp41 gene under conditions described previously ( ).

    Techniques: Knock-Out, Recombinant, Transfection

    Immunoelectron microscopy analysis of Sf9 cells transfected with vAc54KO or vAcWT bacmid DNA at 72 h p.t. using antisera against different nucleocapsid structural proteins. For the experiments shown in panels A to C, Sf9 cells were transfected with vAc54KO

    Journal: Journal of Virology

    Article Title: The Autographa californica Multiple Nucleopolyhedrovirus ac54 Gene Is Crucial for Localization of the Major Capsid Protein VP39 at the Site of Nucleocapsid Assembly

    doi: 10.1128/JVI.02885-15

    Figure Lengend Snippet: Immunoelectron microscopy analysis of Sf9 cells transfected with vAc54KO or vAcWT bacmid DNA at 72 h p.t. using antisera against different nucleocapsid structural proteins. For the experiments shown in panels A to C, Sf9 cells were transfected with vAc54KO

    Article Snippet: To eliminate input bacmid DNA, 1 μg of DNA was digested with 20 units of the DpnI restriction enzyme (NEB) overnight in a 40-μl reaction volume. qPCR was performed using Hot Start PCR master mix III (Chaoshi-Bio) and primers targeting a 100-bp region of the gp41 gene under conditions described previously ( ).

    Techniques: Immuno-Electron Microscopy, Transfection