1 kb dna marker  (New England Biolabs)


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    Name:
    1 kb Plus DNA Ladder
    Description:
    2 Log DNA Ladder 500 1000 gel lanes
    Catalog Number:
    n3200l
    Price:
    231
    Category:
    DNA Ladders
    Size:
    1000 gel lanes
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    New England Biolabs 1 kb dna marker
    1 kb Plus DNA Ladder
    2 Log DNA Ladder 500 1000 gel lanes
    https://www.bioz.com/result/1 kb dna marker/product/New England Biolabs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    1 kb dna marker - by Bioz Stars, 2021-03
    94/100 stars

    Images

    1) Product Images from "Biochemical characterization and chemical validation of Leishmania MAP Kinase-3 as a potential drug target"

    Article Title: Biochemical characterization and chemical validation of Leishmania MAP Kinase-3 as a potential drug target

    Journal: Scientific Reports

    doi: 10.1038/s41598-019-52774-6

    Confirmation of pET28a- Ld MAPK3 construct. Lane 1 showing the amplification of Ld MAPK3 by colony PCR method; Lane 2 is indicating 1-kb DNA ladder and Lane 3 is showing the release of insert ( Ld MAPK3) at 1.2 kb and 5.369 kb as vector (pET 28a(+)) using restriction digestion.
    Figure Legend Snippet: Confirmation of pET28a- Ld MAPK3 construct. Lane 1 showing the amplification of Ld MAPK3 by colony PCR method; Lane 2 is indicating 1-kb DNA ladder and Lane 3 is showing the release of insert ( Ld MAPK3) at 1.2 kb and 5.369 kb as vector (pET 28a(+)) using restriction digestion.

    Techniques Used: Construct, Amplification, Polymerase Chain Reaction, Plasmid Preparation, Positron Emission Tomography

    2) Product Images from "Argonaute of the archaeon Pyrococcus furiosus is a DNA-guided nuclease that targets cognate DNA"

    Article Title: Argonaute of the archaeon Pyrococcus furiosus is a DNA-guided nuclease that targets cognate DNA

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkv415

    Plasmid cleavage by Pf Ago. ( A ) pWUR790 expression plasmid. ( B ) Pf Ago expressed at 20°C and purified in absence of Mn 2+ cleaves expression plasmid pWUR790. Agarose gels with plasmid targets incubated without protein (lane 1), with Pf AgoDM (lane 2) and with Pf Ago (lane 3). M1: 1-kb DNA ladder (New England Biolabs). M2: pWUR790 marker with open circular (OC), linearized (LIN) and supercoiled (SC) pWUR790. ( C ) pWUR704 target plasmid, target site indicated in gray. ( D ) Target region (gray) and FW and RV siDNA guides (black). Predicted cleavage sites are indicated with a black triangle. ( E ) Agarose gels with plasmid targets incubated with Pf AgoDM (lane 1), with guide free Pf Ago (lane 2) and with Pf Ago loaded with FW siDNA, RV siDNA, or both (lane 3–5) in reaction buffer with 250 mM NaCl (left panel) or 500 mM NaCl (right panel). M1: 1 kb GeneRuler marker (Thermo Scientific). M2: pWUR704 marker with open circular (OC), linearized (LIN) and supercoiled (SC) pWUR704.
    Figure Legend Snippet: Plasmid cleavage by Pf Ago. ( A ) pWUR790 expression plasmid. ( B ) Pf Ago expressed at 20°C and purified in absence of Mn 2+ cleaves expression plasmid pWUR790. Agarose gels with plasmid targets incubated without protein (lane 1), with Pf AgoDM (lane 2) and with Pf Ago (lane 3). M1: 1-kb DNA ladder (New England Biolabs). M2: pWUR790 marker with open circular (OC), linearized (LIN) and supercoiled (SC) pWUR790. ( C ) pWUR704 target plasmid, target site indicated in gray. ( D ) Target region (gray) and FW and RV siDNA guides (black). Predicted cleavage sites are indicated with a black triangle. ( E ) Agarose gels with plasmid targets incubated with Pf AgoDM (lane 1), with guide free Pf Ago (lane 2) and with Pf Ago loaded with FW siDNA, RV siDNA, or both (lane 3–5) in reaction buffer with 250 mM NaCl (left panel) or 500 mM NaCl (right panel). M1: 1 kb GeneRuler marker (Thermo Scientific). M2: pWUR704 marker with open circular (OC), linearized (LIN) and supercoiled (SC) pWUR704.

    Techniques Used: Plasmid Preparation, Expressing, Purification, Incubation, Marker

    3) Product Images from "Direct observation of single flexible polymers using single stranded DNA"

    Article Title: Direct observation of single flexible polymers using single stranded DNA

    Journal: Soft matter

    doi: 10.1039/C1SM05297G

    Direct visualization of fluorescently labelled ssDNA molecules using fluorescence microscopy. Single molecules of ssDNA and ds-λ-DNA are shown for (a) stretched and (b) coiled configurations. ssDNA (Sequence 1) with variable dye-labelling ratios are imaged.
    Figure Legend Snippet: Direct visualization of fluorescently labelled ssDNA molecules using fluorescence microscopy. Single molecules of ssDNA and ds-λ-DNA are shown for (a) stretched and (b) coiled configurations. ssDNA (Sequence 1) with variable dye-labelling ratios are imaged.

    Techniques Used: Fluorescence, Microscopy, Sequencing

    4) Product Images from "A selectable, plasmid-based system to generate CRISPR/Cas9 gene edited and knock-in mosquito cell lines"

    Article Title: A selectable, plasmid-based system to generate CRISPR/Cas9 gene edited and knock-in mosquito cell lines

    Journal: Scientific Reports

    doi: 10.1038/s41598-020-80436-5

    CRISPR/Cas9-mediated editing and knock-in of AGO1 in Aag2 cells. ( a ) Three sgRNAs (sg) were designed near the AGO1 translation start site in Aag2 cells. PCRs were designed for surveyor assay (green arrows 1 and 2) and to assess integration (green arrows 1 + and 3). ( b ) Homology-directed repair (HDR) donor template design. The PUb promoter drives expression of red fluorescent protein (RFP); the PUb-RFP cassette is flanked by two loxP sites, followed by a 3xFLAG-tag. Aag2 AGO1 sgRNAs transfections were performed as in Fig. 3 b with the HDR donor template added. Workflows to obtain edited or integrated clones are shown. ( c ) Editing efficiency was assessed by surveyor assay. Expected size of wild-type (WT) amplicon = ~ 410 base pairs (bp; black arrow); expected size of digested fragments based on sgRNA cleavage sites = ~ 180 bp + ~ 230 bp (red arrow); m = marker. ( d ) Single cell clones were sequenced to determine the percentage of edited clones. Immunoblot of AGO1 (top) showed clones with WT and reduced AGO1 protein levels (salmon arrows); A, B, C denote clones. 3xFLAG-tagged Aag2 AGO1 short and long isoforms were detected by both the anti-AGO1 antibody and the anti-FLAG antibody (bottom). ( e ) RFP-positive Aag2 clones from ( b ) were screened for HDR-mediated integration of the donor template. Expected size of amplicons from WT clones = ~ 1500 bp (black arrow); expected size of amplicons from clones containing the integrated HDR donor template = ~ 3700 bp (dark red arrow). B2 and C10 clones were knock-in at the AGO1 locus . ( f ) The PUb-RFP cassette was excised from Aag2 AGO1 knock-in clones (B2 and C10) by transfection of PUb-driven Cre-T2A-pAc and puro selection. WT and un-transfected B2 cells are shown; expected WT and integrated HDR donor template amplicon sizes as in ( e ); expected size of Cre excised amplicon = ~ 1500 bp (black arrow). Puro treatment increases the proportion of Cre excision (B2 + Cre + puro). ( g ) B2 and C10 Cre excised cell lines were sorted for RFP negative clones to obtain homozygous knock-in 3xFLAG- AGO1 Aag2 cell lines. Expected amplicon sizes as in ( e – f ). All full-length immunoblots in Supplementary Fig. 4 ; all full-length DNA gels in Supplementary Fig. 5 .
    Figure Legend Snippet: CRISPR/Cas9-mediated editing and knock-in of AGO1 in Aag2 cells. ( a ) Three sgRNAs (sg) were designed near the AGO1 translation start site in Aag2 cells. PCRs were designed for surveyor assay (green arrows 1 and 2) and to assess integration (green arrows 1 + and 3). ( b ) Homology-directed repair (HDR) donor template design. The PUb promoter drives expression of red fluorescent protein (RFP); the PUb-RFP cassette is flanked by two loxP sites, followed by a 3xFLAG-tag. Aag2 AGO1 sgRNAs transfections were performed as in Fig. 3 b with the HDR donor template added. Workflows to obtain edited or integrated clones are shown. ( c ) Editing efficiency was assessed by surveyor assay. Expected size of wild-type (WT) amplicon = ~ 410 base pairs (bp; black arrow); expected size of digested fragments based on sgRNA cleavage sites = ~ 180 bp + ~ 230 bp (red arrow); m = marker. ( d ) Single cell clones were sequenced to determine the percentage of edited clones. Immunoblot of AGO1 (top) showed clones with WT and reduced AGO1 protein levels (salmon arrows); A, B, C denote clones. 3xFLAG-tagged Aag2 AGO1 short and long isoforms were detected by both the anti-AGO1 antibody and the anti-FLAG antibody (bottom). ( e ) RFP-positive Aag2 clones from ( b ) were screened for HDR-mediated integration of the donor template. Expected size of amplicons from WT clones = ~ 1500 bp (black arrow); expected size of amplicons from clones containing the integrated HDR donor template = ~ 3700 bp (dark red arrow). B2 and C10 clones were knock-in at the AGO1 locus . ( f ) The PUb-RFP cassette was excised from Aag2 AGO1 knock-in clones (B2 and C10) by transfection of PUb-driven Cre-T2A-pAc and puro selection. WT and un-transfected B2 cells are shown; expected WT and integrated HDR donor template amplicon sizes as in ( e ); expected size of Cre excised amplicon = ~ 1500 bp (black arrow). Puro treatment increases the proportion of Cre excision (B2 + Cre + puro). ( g ) B2 and C10 Cre excised cell lines were sorted for RFP negative clones to obtain homozygous knock-in 3xFLAG- AGO1 Aag2 cell lines. Expected amplicon sizes as in ( e – f ). All full-length immunoblots in Supplementary Fig. 4 ; all full-length DNA gels in Supplementary Fig. 5 .

    Techniques Used: CRISPR, Knock-In, Expressing, Transfection, Clone Assay, Amplification, Marker, Selection, Western Blot

    5) Product Images from "The role of Rif1 in telomere length regulation is separable from its role in origin firing"

    Article Title: The role of Rif1 in telomere length regulation is separable from its role in origin firing

    Journal: eLife

    doi: 10.7554/eLife.58066

    DDK overexpression increases origin activation across the genome. ( A ) The DNA content of the arrested cells in Figure 2A and B was measured using flow cytometry. ( B ) The DNA content of the arrested cells in Figure 2C was measured using flow cytometry. ( C ) Relative copy number for the entire Chromosome VI is shown. WT is in purple, ∆N-dbf4 o/e is in orange, and ∆N-dbf4 Cdc7 o/e is in blue, relating to Figure 2C . ( D ) The relative copy number for 1 kb around the midpoint of each origin was calculated for each strain. WT is in purple, ∆N-dbf4 o/e is in orange, and ∆ N-dbf4 Cdc7 o/e is in blue. For each peak, the WT value was subtracted from the value of the over-expression strain and multiplied by 100. Median and interquartile range are plotted over the distribution, and * indicates a significant difference by one-sided Wilcoxon signed rank test: N-dbf4 o/e (p
    Figure Legend Snippet: DDK overexpression increases origin activation across the genome. ( A ) The DNA content of the arrested cells in Figure 2A and B was measured using flow cytometry. ( B ) The DNA content of the arrested cells in Figure 2C was measured using flow cytometry. ( C ) Relative copy number for the entire Chromosome VI is shown. WT is in purple, ∆N-dbf4 o/e is in orange, and ∆N-dbf4 Cdc7 o/e is in blue, relating to Figure 2C . ( D ) The relative copy number for 1 kb around the midpoint of each origin was calculated for each strain. WT is in purple, ∆N-dbf4 o/e is in orange, and ∆ N-dbf4 Cdc7 o/e is in blue. For each peak, the WT value was subtracted from the value of the over-expression strain and multiplied by 100. Median and interquartile range are plotted over the distribution, and * indicates a significant difference by one-sided Wilcoxon signed rank test: N-dbf4 o/e (p

    Techniques Used: Over Expression, Activation Assay, Flow Cytometry

    Mutations in Rif1 that increase telomere length do not affect origin firing across the genome. ( A ) The DNA content of the arrested cells in Figure 4C was measured using flow cytometry. ( B ) The DNA content of the arrested cells in Figure 4D was measured using flow cytometry. ( C ) Relative copy number for the entire Chromosome VI is shown. WT is in purple, rif1-∆1322 is in orange, and rif1 HOOK is in blue, relating to Figure 4D . ( D ) The relative copy number for 1 kb around the midpoint of each origin was calculated for each strain. For each peak, the WT value was subtracted from RIF1 mutant value and multiplied by 100. Median and interquartile range are plotted over the distribution, and one-sided Wilcoxon signed rank test used to test significance: rif1-∆1322 (p=1) and rif1 HOOK (p=1). ( E ) Heatmap of relative copy number for 10 kb region centered at each OriDB origin. Orange indicates high copy number and blue indicates low copy number. Origins are sorted based on highest copy number to lowest copy number.
    Figure Legend Snippet: Mutations in Rif1 that increase telomere length do not affect origin firing across the genome. ( A ) The DNA content of the arrested cells in Figure 4C was measured using flow cytometry. ( B ) The DNA content of the arrested cells in Figure 4D was measured using flow cytometry. ( C ) Relative copy number for the entire Chromosome VI is shown. WT is in purple, rif1-∆1322 is in orange, and rif1 HOOK is in blue, relating to Figure 4D . ( D ) The relative copy number for 1 kb around the midpoint of each origin was calculated for each strain. For each peak, the WT value was subtracted from RIF1 mutant value and multiplied by 100. Median and interquartile range are plotted over the distribution, and one-sided Wilcoxon signed rank test used to test significance: rif1-∆1322 (p=1) and rif1 HOOK (p=1). ( E ) Heatmap of relative copy number for 10 kb region centered at each OriDB origin. Orange indicates high copy number and blue indicates low copy number. Origins are sorted based on highest copy number to lowest copy number.

    Techniques Used: Flow Cytometry, Mutagenesis, Low Copy Number

    6) Product Images from "Using weapons instead of perfume – chemical association strategies of the myrmecophilous bug Scolopostethus pacificus (Rhyparochromidae)"

    Article Title: Using weapons instead of perfume – chemical association strategies of the myrmecophilous bug Scolopostethus pacificus (Rhyparochromidae)

    Journal: bioRxiv

    doi: 10.1101/2020.12.08.412577

    (A) Molecular gut content analysis of Scolopostethus pacificus . Standard ITS2 primers (CAS5ps+CAS28s) amplified the 563bp long bug ITS2, but no ITS amplification was detected with ant-specific Loc1 and Loc2 primers. (B) As a positive control the ant-specific primers Loc1 (167 bp) and Loc2 (228 bp) were amplified with ant DNA. To demonstrate that Loc1 and Loc2 can amplify ant DNA from a predator which recently consumed ants, I dissected guts of Platyusa sonomea rove beetles, that had preyed on ants. For all give rove beetle samples, Loc1 and Loc2 amplified and their identities were confirmed with Sanger sequencing. The ladder shown on the left (A+B) is the 1 kb DNA Ladder from New England Biolabs.
    Figure Legend Snippet: (A) Molecular gut content analysis of Scolopostethus pacificus . Standard ITS2 primers (CAS5ps+CAS28s) amplified the 563bp long bug ITS2, but no ITS amplification was detected with ant-specific Loc1 and Loc2 primers. (B) As a positive control the ant-specific primers Loc1 (167 bp) and Loc2 (228 bp) were amplified with ant DNA. To demonstrate that Loc1 and Loc2 can amplify ant DNA from a predator which recently consumed ants, I dissected guts of Platyusa sonomea rove beetles, that had preyed on ants. For all give rove beetle samples, Loc1 and Loc2 amplified and their identities were confirmed with Sanger sequencing. The ladder shown on the left (A+B) is the 1 kb DNA Ladder from New England Biolabs.

    Techniques Used: Amplification, Positive Control, Sequencing

    7) Product Images from "Single Molecule Hydrodynamic Separation Allows Sensitive and Quantitative Analysis of DNA Conformation and Binding Interactions in Free Solution"

    Article Title: Single Molecule Hydrodynamic Separation Allows Sensitive and Quantitative Analysis of DNA Conformation and Binding Interactions in Free Solution

    Journal: Journal of the American Chemical Society

    doi: 10.1021/jacs.5b10983

    The effects of both sodium chloride (blue) and magnesium chloride (red) on the packing density of double stranded and single stranded DNA is probed by comparing their relative mobilities in the same 1.6 μm diameter capillary. (a) HindIII digested
    Figure Legend Snippet: The effects of both sodium chloride (blue) and magnesium chloride (red) on the packing density of double stranded and single stranded DNA is probed by comparing their relative mobilities in the same 1.6 μm diameter capillary. (a) HindIII digested

    Techniques Used:

    8) Product Images from "The role of Rif1 in telomere length regulation is separable from its role in origin firing"

    Article Title: The role of Rif1 in telomere length regulation is separable from its role in origin firing

    Journal: eLife

    doi: 10.7554/eLife.58066

    DDK overexpression increases origin activation across the genome. ( A ) The DNA content of the arrested cells in Figure 2A and B was measured using flow cytometry. ( B ) The DNA content of the arrested cells in Figure 2C was measured using flow cytometry. ( C ) Relative copy number for the entire Chromosome VI is shown. WT is in purple, ∆N-dbf4 o/e is in orange, and ∆N-dbf4 Cdc7 o/e is in blue, relating to Figure 2C . ( D ) The relative copy number for 1 kb around the midpoint of each origin was calculated for each strain. WT is in purple, ∆N-dbf4 o/e is in orange, and ∆ N-dbf4 Cdc7 o/e is in blue. For each peak, the WT value was subtracted from the value of the over-expression strain and multiplied by 100. Median and interquartile range are plotted over the distribution, and * indicates a significant difference by one-sided Wilcoxon signed rank test: N-dbf4 o/e (p
    Figure Legend Snippet: DDK overexpression increases origin activation across the genome. ( A ) The DNA content of the arrested cells in Figure 2A and B was measured using flow cytometry. ( B ) The DNA content of the arrested cells in Figure 2C was measured using flow cytometry. ( C ) Relative copy number for the entire Chromosome VI is shown. WT is in purple, ∆N-dbf4 o/e is in orange, and ∆N-dbf4 Cdc7 o/e is in blue, relating to Figure 2C . ( D ) The relative copy number for 1 kb around the midpoint of each origin was calculated for each strain. WT is in purple, ∆N-dbf4 o/e is in orange, and ∆ N-dbf4 Cdc7 o/e is in blue. For each peak, the WT value was subtracted from the value of the over-expression strain and multiplied by 100. Median and interquartile range are plotted over the distribution, and * indicates a significant difference by one-sided Wilcoxon signed rank test: N-dbf4 o/e (p

    Techniques Used: Over Expression, Activation Assay, Flow Cytometry

    Mutations in Rif1 that increase telomere length do not affect origin firing across the genome. ( A ) The DNA content of the arrested cells in Figure 4C was measured using flow cytometry. ( B ) The DNA content of the arrested cells in Figure 4D was measured using flow cytometry. ( C ) Relative copy number for the entire Chromosome VI is shown. WT is in purple, rif1-∆1322 is in orange, and rif1 HOOK is in blue, relating to Figure 4D . ( D ) The relative copy number for 1 kb around the midpoint of each origin was calculated for each strain. For each peak, the WT value was subtracted from RIF1 mutant value and multiplied by 100. Median and interquartile range are plotted over the distribution, and one-sided Wilcoxon signed rank test used to test significance: rif1-∆1322 (p=1) and rif1 HOOK (p=1). ( E ) Heatmap of relative copy number for 10 kb region centered at each OriDB origin. Orange indicates high copy number and blue indicates low copy number. Origins are sorted based on highest copy number to lowest copy number.
    Figure Legend Snippet: Mutations in Rif1 that increase telomere length do not affect origin firing across the genome. ( A ) The DNA content of the arrested cells in Figure 4C was measured using flow cytometry. ( B ) The DNA content of the arrested cells in Figure 4D was measured using flow cytometry. ( C ) Relative copy number for the entire Chromosome VI is shown. WT is in purple, rif1-∆1322 is in orange, and rif1 HOOK is in blue, relating to Figure 4D . ( D ) The relative copy number for 1 kb around the midpoint of each origin was calculated for each strain. For each peak, the WT value was subtracted from RIF1 mutant value and multiplied by 100. Median and interquartile range are plotted over the distribution, and one-sided Wilcoxon signed rank test used to test significance: rif1-∆1322 (p=1) and rif1 HOOK (p=1). ( E ) Heatmap of relative copy number for 10 kb region centered at each OriDB origin. Orange indicates high copy number and blue indicates low copy number. Origins are sorted based on highest copy number to lowest copy number.

    Techniques Used: Flow Cytometry, Mutagenesis, Low Copy Number

    9) Product Images from "Practical utilization of recombinant AAV vector reference standards: focus on vector genomes titration by free ITR qPCR"

    Article Title: Practical utilization of recombinant AAV vector reference standards: focus on vector genomes titration by free ITR qPCR

    Journal: Molecular Therapy. Methods & Clinical Development

    doi: 10.1038/mtm.2016.19

    Generation of free ends for the plasmid inverted terminal repeats (ITRs). ( a ) Schematic representation of the plasmid psub201 and the PvuII and HindIII restriction sites. ( b ) Magnification of the plasmid DNA sequences close to the PvuII digestion sites. pEMBL8(+) plasmid backbone (lower case); AAV2-sub201 viral genome (upper case) and PvuII site (CAG/CTG): underlined. ( c ) Separation of undigested and digested plasmid DNA on a 1% agarose gel; supercoiled and linear DNA ladder were used as electrophoresis standards. ( d ) Plasmid DNA purity and concentration measured by spectrophotometry.
    Figure Legend Snippet: Generation of free ends for the plasmid inverted terminal repeats (ITRs). ( a ) Schematic representation of the plasmid psub201 and the PvuII and HindIII restriction sites. ( b ) Magnification of the plasmid DNA sequences close to the PvuII digestion sites. pEMBL8(+) plasmid backbone (lower case); AAV2-sub201 viral genome (upper case) and PvuII site (CAG/CTG): underlined. ( c ) Separation of undigested and digested plasmid DNA on a 1% agarose gel; supercoiled and linear DNA ladder were used as electrophoresis standards. ( d ) Plasmid DNA purity and concentration measured by spectrophotometry.

    Techniques Used: Plasmid Preparation, CTG Assay, Agarose Gel Electrophoresis, Electrophoresis, Concentration Assay, Spectrophotometry

    Related Articles

    Marker:

    Article Title: Argonaute of the archaeon Pyrococcus furiosus is a DNA-guided nuclease that targets cognate DNA
    Article Snippet: Samples were mixed with 6× loading dye (Thermo Scientific) before they were resolved on 0.8% agarose gels. .. As marker, either a 1 kb Generuler Marker (Thermo Scientific) or 1 kb DNA ladder (New England Biolabs) and additionally a custom plasmid marker, were used. .. The custom plasmid marker consisted of non-treated pWUR704 (mostly in supercoiled conformation), Nb.BSMI (New England Biolabs) nicked pWUR704 (open circular conformation) and BcuI (Thermo Scientific) linearized pWUR704.

    Article Title: Direct observation of single flexible polymers using single stranded DNA
    Article Snippet: .. To assay ssDNA products from RCR reactions, agarose gels (0.6%) were run in TAE buffer using 1 kb DNA ladder (NEB) supplemented with 50 ng of λ-DNA as a standard marker. ..

    Article Title: Biochemical characterization and chemical validation of Leishmania MAP Kinase-3 as a potential drug target
    Article Snippet: The enhanced chemiluminescence (ECL) reagent substrate was purchased from BIO-RAD. .. Restriction enzymes, 1-kb DNA marker, stained and unstained protein markers were purchased from NEB. .. Cloning of recombinant construct The expression construct for the production of recombinant MAPK3 was created by using the gene encoding Ld MAPK3 from the NCBI nucleotide database with accession number (XM_003858842.1).

    Plasmid Preparation:

    Article Title: Argonaute of the archaeon Pyrococcus furiosus is a DNA-guided nuclease that targets cognate DNA
    Article Snippet: Samples were mixed with 6× loading dye (Thermo Scientific) before they were resolved on 0.8% agarose gels. .. As marker, either a 1 kb Generuler Marker (Thermo Scientific) or 1 kb DNA ladder (New England Biolabs) and additionally a custom plasmid marker, were used. .. The custom plasmid marker consisted of non-treated pWUR704 (mostly in supercoiled conformation), Nb.BSMI (New England Biolabs) nicked pWUR704 (open circular conformation) and BcuI (Thermo Scientific) linearized pWUR704.

    Amplification:

    Article Title: Using weapons instead of perfume – chemical association strategies of the myrmecophilous bug Scolopostethus pacificus (Rhyparochromidae)
    Article Snippet: DNA extracts of the specimens were used in PCR reactions (initial denaturation 2 min at 95°C; 35 cycles of 0.5 min at 95°C; 1 min at 58°C, 1 min at 72°C; final extension for 5 min at 72°C and final indefinite hold at 4°C) using ITS2 primers [ supplementary Table S1 ; ] with GoTaq Green (12.5 µl master mix, 8.5 µl nuclease-free water, 2 ul 10 µM mixture of F/R primer, 2 µl template). .. A volume (5 µL) of the reaction was then checked for successful amplification on a 1% agarose with the 1 kb DNA Ladder from New England Biolabs and afterwards the remaining PCR product was purified using the Monarch® PCR & DNA Cleanup Kit (New England Biolabs; Ipswich, MA). .. Purified products for S. pacificus and L. occidentale were sent for Sanger sequencing with Laragen Inc. (Culver City, CA).

    Polymerase Chain Reaction:

    Article Title: Using weapons instead of perfume – chemical association strategies of the myrmecophilous bug Scolopostethus pacificus (Rhyparochromidae)
    Article Snippet: DNA extracts of the specimens were used in PCR reactions (initial denaturation 2 min at 95°C; 35 cycles of 0.5 min at 95°C; 1 min at 58°C, 1 min at 72°C; final extension for 5 min at 72°C and final indefinite hold at 4°C) using ITS2 primers [ supplementary Table S1 ; ] with GoTaq Green (12.5 µl master mix, 8.5 µl nuclease-free water, 2 ul 10 µM mixture of F/R primer, 2 µl template). .. A volume (5 µL) of the reaction was then checked for successful amplification on a 1% agarose with the 1 kb DNA Ladder from New England Biolabs and afterwards the remaining PCR product was purified using the Monarch® PCR & DNA Cleanup Kit (New England Biolabs; Ipswich, MA). .. Purified products for S. pacificus and L. occidentale were sent for Sanger sequencing with Laragen Inc. (Culver City, CA).

    Article Title: A selectable, plasmid-based system to generate CRISPR/Cas9 gene edited and knock-in mosquito cell lines
    Article Snippet: All sorting was performed using a BD FACSAria II sorter (BD Biosciences) with a 100 µm nozzle and a sheath pressure of ~ 20 lbf/in2 into 96-well plates containing 50% fresh media, and 50% conditioned media. .. Cells were expanded and genomic DNA was isolated and screened for integration or excision by PCR (primers RU-O-26075 and RU-O-26076); amplicons were visualized on ~ 1% agarose gels with 1 kb plus DNA ladder (NEB) and SYBR Gold (Thermo Fisher Scientific). ..

    Article Title: The role of Rif1 in telomere length regulation is separable from its role in origin firing
    Article Snippet: The DNA was then vacuum transferred onto Hybond+ Nylon (GE Healthcare RPN303B) in 10XSSC, and blocked in Church buffer at 65°C. .. A 32 P radiolabelled Y’ PCR fragment (oligo sequences in ) or 2-log ladder (NEB N3200L) was added at 106 counts/ml of Y’ and 104 counts/ml of 2-log ladder and hybridized overnight. .. The Southern was washed with 1XSSC + 0.1% SDS and imaged using a Storm 825 phosphorimager (GE Healthcare) usually after overnight exposure and analyzed with ImageQuant software.

    Purification:

    Article Title: Using weapons instead of perfume – chemical association strategies of the myrmecophilous bug Scolopostethus pacificus (Rhyparochromidae)
    Article Snippet: DNA extracts of the specimens were used in PCR reactions (initial denaturation 2 min at 95°C; 35 cycles of 0.5 min at 95°C; 1 min at 58°C, 1 min at 72°C; final extension for 5 min at 72°C and final indefinite hold at 4°C) using ITS2 primers [ supplementary Table S1 ; ] with GoTaq Green (12.5 µl master mix, 8.5 µl nuclease-free water, 2 ul 10 µM mixture of F/R primer, 2 µl template). .. A volume (5 µL) of the reaction was then checked for successful amplification on a 1% agarose with the 1 kb DNA Ladder from New England Biolabs and afterwards the remaining PCR product was purified using the Monarch® PCR & DNA Cleanup Kit (New England Biolabs; Ipswich, MA). .. Purified products for S. pacificus and L. occidentale were sent for Sanger sequencing with Laragen Inc. (Culver City, CA).

    Lambda DNA Preparation:

    Article Title: Single Molecule Hydrodynamic Separation Allows Sensitive and Quantitative Analysis of DNA Conformation and Binding Interactions in Free Solution
    Article Snippet: .. Lambda DNA, HindIII digested Lambda DNA, 1 kb DNA Ladder, and Supercoiled DNA Ladder (all from New England Biolabs, Inc.) were used as double stranded DNA samples. .. Staining was performed at 5 or 10 ng/μL total dsDNA concentration and 1 μM TOTO-3 Iodide (Life Technologies) for at least 1 hour in the dark.

    Agarose Gel Electrophoresis:

    Article Title: The role of Rif1 in telomere length regulation is separable from its role in origin firing
    Article Snippet: For restriction digestion, we cut 10 µl of gDNA with XhoI (NEB R0146) to visualize Y’ telomere fragments. .. We loaded genomic digests and 100 ng of 2-log ladder (NEB N3200) onto a 1% agarose gel and subjected it to electrophoresis in 1XTTE overnight at 49V. .. The DNA was then vacuum transferred onto Hybond+ Nylon (GE Healthcare RPN303B) in 10XSSC, and blocked in Church buffer at 65°C.

    Electrophoresis:

    Article Title: The role of Rif1 in telomere length regulation is separable from its role in origin firing
    Article Snippet: For restriction digestion, we cut 10 µl of gDNA with XhoI (NEB R0146) to visualize Y’ telomere fragments. .. We loaded genomic digests and 100 ng of 2-log ladder (NEB N3200) onto a 1% agarose gel and subjected it to electrophoresis in 1XTTE overnight at 49V. .. The DNA was then vacuum transferred onto Hybond+ Nylon (GE Healthcare RPN303B) in 10XSSC, and blocked in Church buffer at 65°C.

    Isolation:

    Article Title: A selectable, plasmid-based system to generate CRISPR/Cas9 gene edited and knock-in mosquito cell lines
    Article Snippet: All sorting was performed using a BD FACSAria II sorter (BD Biosciences) with a 100 µm nozzle and a sheath pressure of ~ 20 lbf/in2 into 96-well plates containing 50% fresh media, and 50% conditioned media. .. Cells were expanded and genomic DNA was isolated and screened for integration or excision by PCR (primers RU-O-26075 and RU-O-26076); amplicons were visualized on ~ 1% agarose gels with 1 kb plus DNA ladder (NEB) and SYBR Gold (Thermo Fisher Scientific). ..

    Staining:

    Article Title: Biochemical characterization and chemical validation of Leishmania MAP Kinase-3 as a potential drug target
    Article Snippet: The enhanced chemiluminescence (ECL) reagent substrate was purchased from BIO-RAD. .. Restriction enzymes, 1-kb DNA marker, stained and unstained protein markers were purchased from NEB. .. Cloning of recombinant construct The expression construct for the production of recombinant MAPK3 was created by using the gene encoding Ld MAPK3 from the NCBI nucleotide database with accession number (XM_003858842.1).

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    New England Biolabs 1 kb dna marker
    Confirmation of pET28a- Ld MAPK3 construct. Lane 1 showing the amplification of Ld MAPK3 by colony PCR method; Lane 2 is indicating <t>1-kb</t> <t>DNA</t> ladder and Lane 3 is showing the release of insert ( Ld MAPK3) at 1.2 kb and 5.369 kb as vector (pET 28a(+)) using restriction digestion.
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    Confirmation of pET28a- Ld MAPK3 construct. Lane 1 showing the amplification of Ld MAPK3 by colony PCR method; Lane 2 is indicating 1-kb DNA ladder and Lane 3 is showing the release of insert ( Ld MAPK3) at 1.2 kb and 5.369 kb as vector (pET 28a(+)) using restriction digestion.

    Journal: Scientific Reports

    Article Title: Biochemical characterization and chemical validation of Leishmania MAP Kinase-3 as a potential drug target

    doi: 10.1038/s41598-019-52774-6

    Figure Lengend Snippet: Confirmation of pET28a- Ld MAPK3 construct. Lane 1 showing the amplification of Ld MAPK3 by colony PCR method; Lane 2 is indicating 1-kb DNA ladder and Lane 3 is showing the release of insert ( Ld MAPK3) at 1.2 kb and 5.369 kb as vector (pET 28a(+)) using restriction digestion.

    Article Snippet: Restriction enzymes, 1-kb DNA marker, stained and unstained protein markers were purchased from NEB.

    Techniques: Construct, Amplification, Polymerase Chain Reaction, Plasmid Preparation, Positron Emission Tomography

    Plasmid cleavage by Pf Ago. ( A ) pWUR790 expression plasmid. ( B ) Pf Ago expressed at 20°C and purified in absence of Mn 2+ cleaves expression plasmid pWUR790. Agarose gels with plasmid targets incubated without protein (lane 1), with Pf AgoDM (lane 2) and with Pf Ago (lane 3). M1: 1-kb DNA ladder (New England Biolabs). M2: pWUR790 marker with open circular (OC), linearized (LIN) and supercoiled (SC) pWUR790. ( C ) pWUR704 target plasmid, target site indicated in gray. ( D ) Target region (gray) and FW and RV siDNA guides (black). Predicted cleavage sites are indicated with a black triangle. ( E ) Agarose gels with plasmid targets incubated with Pf AgoDM (lane 1), with guide free Pf Ago (lane 2) and with Pf Ago loaded with FW siDNA, RV siDNA, or both (lane 3–5) in reaction buffer with 250 mM NaCl (left panel) or 500 mM NaCl (right panel). M1: 1 kb GeneRuler marker (Thermo Scientific). M2: pWUR704 marker with open circular (OC), linearized (LIN) and supercoiled (SC) pWUR704.

    Journal: Nucleic Acids Research

    Article Title: Argonaute of the archaeon Pyrococcus furiosus is a DNA-guided nuclease that targets cognate DNA

    doi: 10.1093/nar/gkv415

    Figure Lengend Snippet: Plasmid cleavage by Pf Ago. ( A ) pWUR790 expression plasmid. ( B ) Pf Ago expressed at 20°C and purified in absence of Mn 2+ cleaves expression plasmid pWUR790. Agarose gels with plasmid targets incubated without protein (lane 1), with Pf AgoDM (lane 2) and with Pf Ago (lane 3). M1: 1-kb DNA ladder (New England Biolabs). M2: pWUR790 marker with open circular (OC), linearized (LIN) and supercoiled (SC) pWUR790. ( C ) pWUR704 target plasmid, target site indicated in gray. ( D ) Target region (gray) and FW and RV siDNA guides (black). Predicted cleavage sites are indicated with a black triangle. ( E ) Agarose gels with plasmid targets incubated with Pf AgoDM (lane 1), with guide free Pf Ago (lane 2) and with Pf Ago loaded with FW siDNA, RV siDNA, or both (lane 3–5) in reaction buffer with 250 mM NaCl (left panel) or 500 mM NaCl (right panel). M1: 1 kb GeneRuler marker (Thermo Scientific). M2: pWUR704 marker with open circular (OC), linearized (LIN) and supercoiled (SC) pWUR704.

    Article Snippet: As marker, either a 1 kb Generuler Marker (Thermo Scientific) or 1 kb DNA ladder (New England Biolabs) and additionally a custom plasmid marker, were used.

    Techniques: Plasmid Preparation, Expressing, Purification, Incubation, Marker

    Direct visualization of fluorescently labelled ssDNA molecules using fluorescence microscopy. Single molecules of ssDNA and ds-λ-DNA are shown for (a) stretched and (b) coiled configurations. ssDNA (Sequence 1) with variable dye-labelling ratios are imaged.

    Journal: Soft matter

    Article Title: Direct observation of single flexible polymers using single stranded DNA

    doi: 10.1039/C1SM05297G

    Figure Lengend Snippet: Direct visualization of fluorescently labelled ssDNA molecules using fluorescence microscopy. Single molecules of ssDNA and ds-λ-DNA are shown for (a) stretched and (b) coiled configurations. ssDNA (Sequence 1) with variable dye-labelling ratios are imaged.

    Article Snippet: To assay ssDNA products from RCR reactions, agarose gels (0.6%) were run in TAE buffer using 1 kb DNA ladder (NEB) supplemented with 50 ng of λ-DNA as a standard marker.

    Techniques: Fluorescence, Microscopy, Sequencing

    CRISPR/Cas9-mediated editing and knock-in of AGO1 in Aag2 cells. ( a ) Three sgRNAs (sg) were designed near the AGO1 translation start site in Aag2 cells. PCRs were designed for surveyor assay (green arrows 1 and 2) and to assess integration (green arrows 1 + and 3). ( b ) Homology-directed repair (HDR) donor template design. The PUb promoter drives expression of red fluorescent protein (RFP); the PUb-RFP cassette is flanked by two loxP sites, followed by a 3xFLAG-tag. Aag2 AGO1 sgRNAs transfections were performed as in Fig. 3 b with the HDR donor template added. Workflows to obtain edited or integrated clones are shown. ( c ) Editing efficiency was assessed by surveyor assay. Expected size of wild-type (WT) amplicon = ~ 410 base pairs (bp; black arrow); expected size of digested fragments based on sgRNA cleavage sites = ~ 180 bp + ~ 230 bp (red arrow); m = marker. ( d ) Single cell clones were sequenced to determine the percentage of edited clones. Immunoblot of AGO1 (top) showed clones with WT and reduced AGO1 protein levels (salmon arrows); A, B, C denote clones. 3xFLAG-tagged Aag2 AGO1 short and long isoforms were detected by both the anti-AGO1 antibody and the anti-FLAG antibody (bottom). ( e ) RFP-positive Aag2 clones from ( b ) were screened for HDR-mediated integration of the donor template. Expected size of amplicons from WT clones = ~ 1500 bp (black arrow); expected size of amplicons from clones containing the integrated HDR donor template = ~ 3700 bp (dark red arrow). B2 and C10 clones were knock-in at the AGO1 locus . ( f ) The PUb-RFP cassette was excised from Aag2 AGO1 knock-in clones (B2 and C10) by transfection of PUb-driven Cre-T2A-pAc and puro selection. WT and un-transfected B2 cells are shown; expected WT and integrated HDR donor template amplicon sizes as in ( e ); expected size of Cre excised amplicon = ~ 1500 bp (black arrow). Puro treatment increases the proportion of Cre excision (B2 + Cre + puro). ( g ) B2 and C10 Cre excised cell lines were sorted for RFP negative clones to obtain homozygous knock-in 3xFLAG- AGO1 Aag2 cell lines. Expected amplicon sizes as in ( e – f ). All full-length immunoblots in Supplementary Fig. 4 ; all full-length DNA gels in Supplementary Fig. 5 .

    Journal: Scientific Reports

    Article Title: A selectable, plasmid-based system to generate CRISPR/Cas9 gene edited and knock-in mosquito cell lines

    doi: 10.1038/s41598-020-80436-5

    Figure Lengend Snippet: CRISPR/Cas9-mediated editing and knock-in of AGO1 in Aag2 cells. ( a ) Three sgRNAs (sg) were designed near the AGO1 translation start site in Aag2 cells. PCRs were designed for surveyor assay (green arrows 1 and 2) and to assess integration (green arrows 1 + and 3). ( b ) Homology-directed repair (HDR) donor template design. The PUb promoter drives expression of red fluorescent protein (RFP); the PUb-RFP cassette is flanked by two loxP sites, followed by a 3xFLAG-tag. Aag2 AGO1 sgRNAs transfections were performed as in Fig. 3 b with the HDR donor template added. Workflows to obtain edited or integrated clones are shown. ( c ) Editing efficiency was assessed by surveyor assay. Expected size of wild-type (WT) amplicon = ~ 410 base pairs (bp; black arrow); expected size of digested fragments based on sgRNA cleavage sites = ~ 180 bp + ~ 230 bp (red arrow); m = marker. ( d ) Single cell clones were sequenced to determine the percentage of edited clones. Immunoblot of AGO1 (top) showed clones with WT and reduced AGO1 protein levels (salmon arrows); A, B, C denote clones. 3xFLAG-tagged Aag2 AGO1 short and long isoforms were detected by both the anti-AGO1 antibody and the anti-FLAG antibody (bottom). ( e ) RFP-positive Aag2 clones from ( b ) were screened for HDR-mediated integration of the donor template. Expected size of amplicons from WT clones = ~ 1500 bp (black arrow); expected size of amplicons from clones containing the integrated HDR donor template = ~ 3700 bp (dark red arrow). B2 and C10 clones were knock-in at the AGO1 locus . ( f ) The PUb-RFP cassette was excised from Aag2 AGO1 knock-in clones (B2 and C10) by transfection of PUb-driven Cre-T2A-pAc and puro selection. WT and un-transfected B2 cells are shown; expected WT and integrated HDR donor template amplicon sizes as in ( e ); expected size of Cre excised amplicon = ~ 1500 bp (black arrow). Puro treatment increases the proportion of Cre excision (B2 + Cre + puro). ( g ) B2 and C10 Cre excised cell lines were sorted for RFP negative clones to obtain homozygous knock-in 3xFLAG- AGO1 Aag2 cell lines. Expected amplicon sizes as in ( e – f ). All full-length immunoblots in Supplementary Fig. 4 ; all full-length DNA gels in Supplementary Fig. 5 .

    Article Snippet: Cells were expanded and genomic DNA was isolated and screened for integration or excision by PCR (primers RU-O-26075 and RU-O-26076); amplicons were visualized on ~ 1% agarose gels with 1 kb plus DNA ladder (NEB) and SYBR Gold (Thermo Fisher Scientific).

    Techniques: CRISPR, Knock-In, Expressing, Transfection, Clone Assay, Amplification, Marker, Selection, Western Blot