molecular weight dna ladder  (New England Biolabs)


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    Low Molecular Weight DNA Ladder
    Description:
    Low Molecular Weight DNA Ladder 500 gel lanes
    Catalog Number:
    n3233l
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    270
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    500 gel lanes
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    DNA Ladders
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    Structured Review

    New England Biolabs molecular weight dna ladder
    Low Molecular Weight DNA Ladder
    Low Molecular Weight DNA Ladder 500 gel lanes
    https://www.bioz.com/result/molecular weight dna ladder/product/New England Biolabs
    Average 95 stars, based on 107 article reviews
    Price from $9.99 to $1999.99
    molecular weight dna ladder - by Bioz Stars, 2020-02
    95/100 stars

    Images

    1) Product Images from "Detection of the pediocin gene pedA in strains from human faeces by real-time PCR and characterization of Pediococcus acidilactici UVA1"

    Article Title: Detection of the pediocin gene pedA in strains from human faeces by real-time PCR and characterization of Pediococcus acidilactici UVA1

    Journal: BMC Biotechnology

    doi: 10.1186/1472-6750-7-55

    Transcription analysis of pedA . pedA -reverse transcription-PCR on cDNA from P. acidilactici bac- (1, 2, 3) or UVA1 (7, 8, 9) after 1 h 30, 2 h 30 and 3 h 30 of growth, respectively, and from P. acidilactici DSM 20284 T (4) or UL5 (6) and P. pentosaceus DSM 20336 T (5) after 2 h 30 of growth. 10: water instead of DNA. lm: low molecular weight DNA ladder (in bp). h: Tridye 100-bp DNA ladder (in bp). Expected product size: 100 bp.
    Figure Legend Snippet: Transcription analysis of pedA . pedA -reverse transcription-PCR on cDNA from P. acidilactici bac- (1, 2, 3) or UVA1 (7, 8, 9) after 1 h 30, 2 h 30 and 3 h 30 of growth, respectively, and from P. acidilactici DSM 20284 T (4) or UL5 (6) and P. pentosaceus DSM 20336 T (5) after 2 h 30 of growth. 10: water instead of DNA. lm: low molecular weight DNA ladder (in bp). h: Tridye 100-bp DNA ladder (in bp). Expected product size: 100 bp.

    Techniques Used: Polymerase Chain Reaction, BAC Assay, Molecular Weight

    2) Product Images from "RIPiT-Seq: A high-throughput approach for footprinting RNA:protein complexes"

    Article Title: RIPiT-Seq: A high-throughput approach for footprinting RNA:protein complexes

    Journal: Methods (San Diego, Calif.)

    doi: 10.1016/j.ymeth.2013.09.013

    UV-crosslinking bias of different RBPs dictates the suitability of different approaches for identifying binding sites A. hnRNPA1:ssDNA interface. Crystal structure of two-RRM-containing UP1 domain of hnRNP A1 (PDB ID: 2UP1; blue) in complex with a target containing its AGGG preferred recognition motif (in this case, within single-stranded DNA, ssDNA; red). Black box. Enlarged view of the DNA:protein interface. Aromatic residues (Phe-17 and Phe-59 from RRM1) that stack with the nucleobases are shown in cyan. B. eIF4AIII:RNA interface. Crystal structure of eIF4AIII (PDB ID: 2J0S; blue) complexed with RNA (red) and AMP-PNP. Black box. Enlarged view of the RNA:protein interface. Note that the RNA bases are pointing away from the bound protein. ]. D. RIPiT and CLIP yield different types of information. Top: Two similar yet compositionally distinct hypothetical multi-subunit RNPs. RBPs (blue), non-RBPs (green) and proteins unique to each complex are shown (complex A: yellow; complex B: red). Left: RIPiT can reveal the binding sites of an intact multi-subunit RNP, and can also distinguish between footprints of two compositionally similar complexes (schematics on gray background). However, RIPiT does not conclusively define direct RBP-RNA interactions (crossed-out schematic). Right: On the contrary, while CLIP reveals no information regarding the complexes an RNA-bound RBP is part of (crossed-out schematics), it can unveil the sites of direct contact between an RBP and RNA (bottom schematic).
    Figure Legend Snippet: UV-crosslinking bias of different RBPs dictates the suitability of different approaches for identifying binding sites A. hnRNPA1:ssDNA interface. Crystal structure of two-RRM-containing UP1 domain of hnRNP A1 (PDB ID: 2UP1; blue) in complex with a target containing its AGGG preferred recognition motif (in this case, within single-stranded DNA, ssDNA; red). Black box. Enlarged view of the DNA:protein interface. Aromatic residues (Phe-17 and Phe-59 from RRM1) that stack with the nucleobases are shown in cyan. B. eIF4AIII:RNA interface. Crystal structure of eIF4AIII (PDB ID: 2J0S; blue) complexed with RNA (red) and AMP-PNP. Black box. Enlarged view of the RNA:protein interface. Note that the RNA bases are pointing away from the bound protein. ]. D. RIPiT and CLIP yield different types of information. Top: Two similar yet compositionally distinct hypothetical multi-subunit RNPs. RBPs (blue), non-RBPs (green) and proteins unique to each complex are shown (complex A: yellow; complex B: red). Left: RIPiT can reveal the binding sites of an intact multi-subunit RNP, and can also distinguish between footprints of two compositionally similar complexes (schematics on gray background). However, RIPiT does not conclusively define direct RBP-RNA interactions (crossed-out schematic). Right: On the contrary, while CLIP reveals no information regarding the complexes an RNA-bound RBP is part of (crossed-out schematics), it can unveil the sites of direct contact between an RBP and RNA (bottom schematic).

    Techniques Used: Binding Assay, Cross-linking Immunoprecipitation

    Biochemical analysis of proteins and RNAs from the RIPiT procedure A. Western blots showing tetracycline (Tet)-mediated induction of eIF4AIII protein with the FLAG tag at its N- or C-terminus (top and bottom panels, respectively). The Tet concentration used for induction is indicated at the top of each lane; protein identities are indicated to the right. B. Levels of proteins detected by western blots in different fractions during EJC RIPiT. The table on the top indicates the different fractions from the RIPiT procedure and the antibodies used for 1 st and 2 nd IPs. The stably expressed FLAG-tag fusion protein used in each sample is indicated directly above lane. Proteins detected by western blot are indicated to the right. C. Size distribution of EJC footprints upon RNase I titration. An autoradiogram of 26% denaturing PAGE with 5′ [ 32 P]-labeled RNA fragments from base-hydrolysis of poly U 30 oligonucleotide (lane 1) or FLAG-Magoh:eIF4AIII RIPiT (lanes 2–5). RNase I concentrations used are indicated at top of each lane; nucleotide (nt) lengths are to the left. D. Quantification of desired size RNA footprints in RIPiT elution. An autoradiogram of 20% denaturing PAGE with 5′ [ 32 P]-labeled 100 bp NEB DNA ladder (lane 1), low molecular weight ssDNA ladder (lane 2), footprints of RIPiTs indicated on top (lanes 3 and 4) and a 21 nt ssRNA oligo of known specific activity (lane 5). The signal from red rectangles is quantified in comparison to that of the 21 nt oligo to estimate the amount of RIPiT footprints in the indicated size range.
    Figure Legend Snippet: Biochemical analysis of proteins and RNAs from the RIPiT procedure A. Western blots showing tetracycline (Tet)-mediated induction of eIF4AIII protein with the FLAG tag at its N- or C-terminus (top and bottom panels, respectively). The Tet concentration used for induction is indicated at the top of each lane; protein identities are indicated to the right. B. Levels of proteins detected by western blots in different fractions during EJC RIPiT. The table on the top indicates the different fractions from the RIPiT procedure and the antibodies used for 1 st and 2 nd IPs. The stably expressed FLAG-tag fusion protein used in each sample is indicated directly above lane. Proteins detected by western blot are indicated to the right. C. Size distribution of EJC footprints upon RNase I titration. An autoradiogram of 26% denaturing PAGE with 5′ [ 32 P]-labeled RNA fragments from base-hydrolysis of poly U 30 oligonucleotide (lane 1) or FLAG-Magoh:eIF4AIII RIPiT (lanes 2–5). RNase I concentrations used are indicated at top of each lane; nucleotide (nt) lengths are to the left. D. Quantification of desired size RNA footprints in RIPiT elution. An autoradiogram of 20% denaturing PAGE with 5′ [ 32 P]-labeled 100 bp NEB DNA ladder (lane 1), low molecular weight ssDNA ladder (lane 2), footprints of RIPiTs indicated on top (lanes 3 and 4) and a 21 nt ssRNA oligo of known specific activity (lane 5). The signal from red rectangles is quantified in comparison to that of the 21 nt oligo to estimate the amount of RIPiT footprints in the indicated size range.

    Techniques Used: Western Blot, FLAG-tag, Concentration Assay, Stable Transfection, Titration, Polyacrylamide Gel Electrophoresis, Labeling, Molecular Weight, Activity Assay

    3) Product Images from "Switching the activity of Cas12a using guide RNA strand displacement circuits"

    Article Title: Switching the activity of Cas12a using guide RNA strand displacement circuits

    Journal: Nature Communications

    doi: 10.1038/s41467-019-09953-w

    Principle of strand displacement switchable gRNAs. a When the RNA trigger binds the SD gRNA, the 5′ extension domain occluding handle is displaced, thereby restoring the gRNA handle. Binding of Cas12a leads to cleavage of the gRNA, which removes the 5′ extension and creates an active Cas12a-gRNA complex. The domains are labeled as follows: s—separator, h—handle, t—target, X1—toehold. The overall nomenclature follows Zhang et al. 32 : A domain is denoted by a single letter. An upper index x denotes the first x nucleotides of a domain counting from its 5′ end. A lower index x denotes all but the first x nucleotides of a domain counting from its 5′ end. A combination of upper and lower indices includes those nucleotides that are present in both subdomains. A bar above the letter marks the reverse complement of a domain. The red star marks the position where Cas12a cleaves a successfully bound gRNA. b Agarose gel showing cutting of a target DNA by handle-based SD gRNAs with two different target sequences in the absence and presence of trigger RNA (uncut: 1190 bp, cut t1: 357 bp, cut t2: 506 bp). c Denaturing PAGE showing induced gRNA processing due to trigger binding. d Agarose gel showing activation of target cutting by varying amounts of trigger RNA (uncut: 1190 bp, cut: 357 bp). e Transfer function derived from the fraction of cut target by gels as shown in ( d ) ( N = 3, t -distribution two-sided 90% confidence interval). Source data are provided as a Source Data file
    Figure Legend Snippet: Principle of strand displacement switchable gRNAs. a When the RNA trigger binds the SD gRNA, the 5′ extension domain occluding handle is displaced, thereby restoring the gRNA handle. Binding of Cas12a leads to cleavage of the gRNA, which removes the 5′ extension and creates an active Cas12a-gRNA complex. The domains are labeled as follows: s—separator, h—handle, t—target, X1—toehold. The overall nomenclature follows Zhang et al. 32 : A domain is denoted by a single letter. An upper index x denotes the first x nucleotides of a domain counting from its 5′ end. A lower index x denotes all but the first x nucleotides of a domain counting from its 5′ end. A combination of upper and lower indices includes those nucleotides that are present in both subdomains. A bar above the letter marks the reverse complement of a domain. The red star marks the position where Cas12a cleaves a successfully bound gRNA. b Agarose gel showing cutting of a target DNA by handle-based SD gRNAs with two different target sequences in the absence and presence of trigger RNA (uncut: 1190 bp, cut t1: 357 bp, cut t2: 506 bp). c Denaturing PAGE showing induced gRNA processing due to trigger binding. d Agarose gel showing activation of target cutting by varying amounts of trigger RNA (uncut: 1190 bp, cut: 357 bp). e Transfer function derived from the fraction of cut target by gels as shown in ( d ) ( N = 3, t -distribution two-sided 90% confidence interval). Source data are provided as a Source Data file

    Techniques Used: Binding Assay, Labeling, Agarose Gel Electrophoresis, Polyacrylamide Gel Electrophoresis, Activation Assay, Derivative Assay

    4) Product Images from "Structural diversity of supercoiled DNA"

    Article Title: Structural diversity of supercoiled DNA

    Journal: Nature Communications

    doi: 10.1038/ncomms9440

    Effect of supercoiling on the structure of minicircle DNA. ( a ) Individual 336 bp minicircle topoisomers were isolated and analysed by polyacrylamide gel electrophoresis in the presence of 10 mM CaCl 2 . Mr: 100 bp DNA ladder, L: minicircle linearized by EcoRV, N: minicircle nicked by Nb.BbvCI. ( b ) Projections of cryo-ET subtomograms of hydrated 336 bp DNA minicircles of the Lk =34 topoisomer. ( c ) Commonly observed shapes were open circle, open figure-8, figure-8, racquet, handcuffs, needle, and rod, each of which are shown in orthogonal views. ( d ) Other shapes observed, especially in the more highly supercoiled topoisomers. ( e ) Shape frequency distribution plot for each topoisomer population (n=number of minicircles analysed). A weighted average for each topoisomer, approximating the average degree of compactness, is denoted by the black triangle. The weighted average was calculated by assigning each conformation a value that increased in line with compactness. Open circles were given a value of 1, open figure-8 s a value of 2, figure-8 s as a value of 3, and so on. The relative fraction of each was subsequently used to determine the average degree of compactness. Lk , Δ Lk and superhelical density (σ) for each topoisomer are shown (see Supplementary Note 1 ).
    Figure Legend Snippet: Effect of supercoiling on the structure of minicircle DNA. ( a ) Individual 336 bp minicircle topoisomers were isolated and analysed by polyacrylamide gel electrophoresis in the presence of 10 mM CaCl 2 . Mr: 100 bp DNA ladder, L: minicircle linearized by EcoRV, N: minicircle nicked by Nb.BbvCI. ( b ) Projections of cryo-ET subtomograms of hydrated 336 bp DNA minicircles of the Lk =34 topoisomer. ( c ) Commonly observed shapes were open circle, open figure-8, figure-8, racquet, handcuffs, needle, and rod, each of which are shown in orthogonal views. ( d ) Other shapes observed, especially in the more highly supercoiled topoisomers. ( e ) Shape frequency distribution plot for each topoisomer population (n=number of minicircles analysed). A weighted average for each topoisomer, approximating the average degree of compactness, is denoted by the black triangle. The weighted average was calculated by assigning each conformation a value that increased in line with compactness. Open circles were given a value of 1, open figure-8 s a value of 2, figure-8 s as a value of 3, and so on. The relative fraction of each was subsequently used to determine the average degree of compactness. Lk , Δ Lk and superhelical density (σ) for each topoisomer are shown (see Supplementary Note 1 ).

    Techniques Used: Isolation, Polyacrylamide Gel Electrophoresis

    5) Product Images from "Intra-tumor heterogeneity of MLH1 promoter methylation revealed by deep single molecule bisulfite sequencing"

    Article Title: Intra-tumor heterogeneity of MLH1 promoter methylation revealed by deep single molecule bisulfite sequencing

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp457

    Comparison of sequencing results from conventional cloning and Sanger sequencing and 454 Life Sciences FLX sequencing of MLH1 promoter PCR products from bisulfite treated DNA of an endometrial cancer and normal blood from Patient #1684. ( A ) Schematic of the MLH1 promoter is presented with arrows indicating the location of PCR primers and vertical lines representing the position of CG dinucleotides. Below the schematic are the results from cloning and bisulfite sequencing 45 molecules from the distal (left) and proximal (right) promoter in each sample (tumor and matched normal blood). Each column represents a CG dinucleotide in the sequence, and corresponds to each vertical line in the promoter schematic. Each row represents a single molecule. The color of the boxes represents the methylation state of each cytosine. Red, methylated; Black, unmethylated. ( B ) The results from the FLX single molecule sequencing of the same samples. The distance of each CpG from the transcription start site (UCSC Human Genome March 2006) is listed from distal to proximal. The Distal Amplicon: −671, −662, −654, −648, −634, −632, −630, −626, −623, −619, −609, −605, −596, −584, −576, −569, −566, −564, −560, −558, −548, −540, −537, −512, −505, −483, −470, −465, −449, −446, −421, −405, −380, −368. The Proximal Amplicon: −341, −325, −318, −286, −280, −249, −240, −226, −209, −202, −192, −190, −184, −165, −154, −139, −70, −47, −23, −15. As an additional point of reference, the translation start site (ATG) is currently annotated at +60 downstream of the transcription start site (position 0).
    Figure Legend Snippet: Comparison of sequencing results from conventional cloning and Sanger sequencing and 454 Life Sciences FLX sequencing of MLH1 promoter PCR products from bisulfite treated DNA of an endometrial cancer and normal blood from Patient #1684. ( A ) Schematic of the MLH1 promoter is presented with arrows indicating the location of PCR primers and vertical lines representing the position of CG dinucleotides. Below the schematic are the results from cloning and bisulfite sequencing 45 molecules from the distal (left) and proximal (right) promoter in each sample (tumor and matched normal blood). Each column represents a CG dinucleotide in the sequence, and corresponds to each vertical line in the promoter schematic. Each row represents a single molecule. The color of the boxes represents the methylation state of each cytosine. Red, methylated; Black, unmethylated. ( B ) The results from the FLX single molecule sequencing of the same samples. The distance of each CpG from the transcription start site (UCSC Human Genome March 2006) is listed from distal to proximal. The Distal Amplicon: −671, −662, −654, −648, −634, −632, −630, −626, −623, −619, −609, −605, −596, −584, −576, −569, −566, −564, −560, −558, −548, −540, −537, −512, −505, −483, −470, −465, −449, −446, −421, −405, −380, −368. The Proximal Amplicon: −341, −325, −318, −286, −280, −249, −240, −226, −209, −202, −192, −190, −184, −165, −154, −139, −70, −47, −23, −15. As an additional point of reference, the translation start site (ATG) is currently annotated at +60 downstream of the transcription start site (position 0).

    Techniques Used: Sequencing, Clone Assay, Polymerase Chain Reaction, Methylation Sequencing, Methylation, Amplification

    Experimental design for deep bisulfite sequencing in individual samples. ( A ) Detection of DNA methylation by sodium bisulfite treatment and PCR. ( B ) PCR amplification with sample specific DNA barcodes. ( C ) Pool PCR products for 454 sequencing.
    Figure Legend Snippet: Experimental design for deep bisulfite sequencing in individual samples. ( A ) Detection of DNA methylation by sodium bisulfite treatment and PCR. ( B ) PCR amplification with sample specific DNA barcodes. ( C ) Pool PCR products for 454 sequencing.

    Techniques Used: Methylation Sequencing, DNA Methylation Assay, Polymerase Chain Reaction, Amplification, Sequencing

    Related Articles

    Amplification:

    Article Title: Differentiated Human Midbrain-Derived Neural Progenitor Cells Express Excitatory Strychnine-Sensitive Glycine Receptors Containing ?2? Subunits
    Article Snippet: .. The correct amplicon size was asserted by agarose gel electrophoresis using low molecular weight DNA ladder (New England Biolabs, Ipswich, MA, USA). .. Oligonucleotide primers for human glycine receptor subunits α1–4 and β ( ) were designed to flank intron sequences, if feasible, using Primer 3 software.

    Synthesized:

    Article Title: Dual coding potential of a 2′,5′-branched ribonucleotide in DNA
    Article Snippet: .. Single-stranded oligonucleotides synthesized by Eurofins MWG Operon or a Low Molecular Weight DNA Ladder (New England BioLabs, NEB) were used as size markers in gel electrophoresis. ..

    Electrophoresis:

    Article Title: A LysR-family transcriptional regulator required for virulence in Brucella abortus is highly conserved among the α-proteobacteria
    Article Snippet: A low-molecular-weight DNA ladder (New England BioLabs, Ipswich, MA, USA) was labeled with [γ-32 P]-ATP (Perkin-Elmer, San Jose, CA, USA) and polynucleotide kinase. .. Following electrophoresis in 1 × TBE buffer, the ladder and RNA samples were transferred to an Amersham Hybond™-N+ membrane (GE Healthcare, Piscataway, NJ, USA) by electroblotting in 1 × TBE buffer.

    Article Title: Endoribonuclease YbeY Is Linked to Proper Cellular Morphology and Virulence in Brucella abortus
    Article Snippet: A low-molecular-weight DNA ladder (New England BioLabs, Ipswich, MA) was labeled with [γ-32 P]ATP and polynucleotide kinase, and this radiolabeled ladder was also separated on the polyacrylamide gel. .. Following electrophoresis in 1× TBE buffer, the ladder and RNA samples were transferred to an Amersham Hybond-N+ membrane (GE Healthcare, Piscataway, NJ) in 1× TBE buffer.

    Incubation:

    Article Title: RNase H1 directs origin-specific initiation of DNA replication in human mitochondria
    Article Snippet: The reactions were incubated at 32°C for 5 min and stopped by the addition of 200 μL stop buffer (10 mM Tris-HCl pH 8.0, 0.2 M NaCl, 1 mM EDTA, 100 μg/mL glycogen (Roche) and 100 μg/mL proteinase K (Ambion)) followed by incubation at 42°C for 45 min. .. Low Molecular Weight DNA Ladder (NEB) was used as a size marker.

    Hybridization:

    Article Title: A LysR-family transcriptional regulator required for virulence in Brucella abortus is highly conserved among the α-proteobacteria
    Article Snippet: A low-molecular-weight DNA ladder (New England BioLabs, Ipswich, MA, USA) was labeled with [γ-32 P]-ATP (Perkin-Elmer, San Jose, CA, USA) and polynucleotide kinase. .. The samples were UV-cross-linked to the membrane and the membrane was pre-hybridized in ULTRAhyb® -Oligo Buffer (Ambion, Austin, TX, USA) for 2 h at ~45°C in a rotating hybridization oven.

    Article Title: Endoribonuclease YbeY Is Linked to Proper Cellular Morphology and Virulence in Brucella abortus
    Article Snippet: A low-molecular-weight DNA ladder (New England BioLabs, Ipswich, MA) was labeled with [γ-32 P]ATP and polynucleotide kinase, and this radiolabeled ladder was also separated on the polyacrylamide gel. .. The samples were UV cross-linked to the membrane, and the membrane was prehybridized in ULTRAhyb-Oligo buffer (Ambion, Austin, TX) for 45 min at ∼42°C in a rotating hybridization oven.

    Northern Blot:

    Article Title: Experimental discovery of sRNAs in Vibrio cholerae by direct cloning, 5S/tRNA depletion and parallel sequencing
    Article Snippet: Generally, northern blots were performed with RNA probes transcribed from PCR-derived templates ( Supplementary Table 4 ) with T7 promoters using 32 P-UTP and T7 polymerase (Promega) according to the manufacturer's instructions. .. Low molecular weight DNA ladder (NEB) was run alongside RNA samples to provide estimations for the sizes of RNA bands.

    Article Title: A LysR-family transcriptional regulator required for virulence in Brucella abortus is highly conserved among the α-proteobacteria
    Article Snippet: Paragraph title: Northern blot analysis ... A low-molecular-weight DNA ladder (New England BioLabs, Ipswich, MA, USA) was labeled with [γ-32 P]-ATP (Perkin-Elmer, San Jose, CA, USA) and polynucleotide kinase.

    Article Title: Sibling sRNA RyfA1 Influences Shigella dysenteriae Pathogenesis
    Article Snippet: Paragraph title: 2.7. Northern Blot ... A Low Molecular Weight DNA Ladder (New England BioLabs, Inc.) was labeled with [ɣ-32P]-ATP using polynucleotide kinase and run on the same gel as the RNA sample.

    Article Title: Endoribonuclease YbeY Is Linked to Proper Cellular Morphology and Virulence in Brucella abortus
    Article Snippet: Paragraph title: Northern blot analysis. ... A low-molecular-weight DNA ladder (New England BioLabs, Ipswich, MA) was labeled with [γ-32 P]ATP and polynucleotide kinase, and this radiolabeled ladder was also separated on the polyacrylamide gel.

    other:

    Article Title: Genome-wide mapping of nucleotide excision repair with XR-seq
    Article Snippet: 34 Load 6 μ l of the mixture for each sample and 2 μ l of low-molecular-weight DNA ladder to the wells of a 10% native polyacrylamide gel, and run the gel in 1× TBE buffer at 125 V for ~2 h. CRITICAL STEP It is important to stop running the gel when the cyan dye reaches the bottom of the gel.

    Article Title: Genome-wide mapping of nucleotide excision repair with XR-seq
    Article Snippet: Load 6 μ l of the mixture from each sample and 2 μ l of low-molecular-weight DNA ladder to the wells of a 10% native polyacrylamide gel, and run the gel in 1× TBE buffer at 125 V for 1.5 h. 29.

    Reverse Transcription Polymerase Chain Reaction:

    Article Title: Blockade of Eosinophil-Induced Bronchial Epithelial-Mesenchymal Transition with a Geranyl Acetophenone in a Coculture Model
    Article Snippet: Paragraph title: RT-PCR ... PCR products in each gel electrophoresis were ran in parallel to a Low Molecular Weight DNA Ladder (NEB, USA).

    Molecular Weight:

    Article Title: Experimental discovery of sRNAs in Vibrio cholerae by direct cloning, 5S/tRNA depletion and parallel sequencing
    Article Snippet: .. Low molecular weight DNA ladder (NEB) was run alongside RNA samples to provide estimations for the sizes of RNA bands. .. Total RNA (10–40 µg) was run on denaturing polyacrylamide gels (10% or 15%) and transferred to Hybond N+ membranes (Amersham) in 1× TBE using the Mini Trans-Blot Cell apparatus (Bio-Rad) according to the manufacturer's instructions.

    Article Title: RNase H1 directs origin-specific initiation of DNA replication in human mitochondria
    Article Snippet: .. Low Molecular Weight DNA Ladder (NEB) was used as a size marker. ..

    Article Title: Sibling sRNA RyfA1 Influences Shigella dysenteriae Pathogenesis
    Article Snippet: .. A Low Molecular Weight DNA Ladder (New England BioLabs, Inc.) was labeled with [ɣ-32P]-ATP using polynucleotide kinase and run on the same gel as the RNA sample. ..

    Article Title: Dual coding potential of a 2′,5′-branched ribonucleotide in DNA
    Article Snippet: .. Single-stranded oligonucleotides synthesized by Eurofins MWG Operon or a Low Molecular Weight DNA Ladder (New England BioLabs, NEB) were used as size markers in gel electrophoresis. ..

    Article Title: Structural diversity of supercoiled DNA
    Article Snippet: .. BbvCI, EcoRV, Nb.BbvCI, NdeI, Nuclease Bal-31, T4 DNA Ligase, low molecular weight DNA ladder and 100 bp DNA ladder were purchased from New England Biolabs (Ipswich, MA). .. Proteinase K was purchased from Roche Molecular Biochemicals (Mannheim, Germany).

    Article Title: Interaction of double-stranded DNA with polymerized PprA protein from Deinococcus radiodurans
    Article Snippet: The low-molecular-weight DNA ladder (New England Biolabs) was composed of 11 DNA fragments (25, 50, 75, 100, 150, 200, 250, 300, 350, 500, and 766 bp). .. SeaKem GTG Agarose and Nusieve 3:1 Agarose (Takara Bio) were used to prepare agarose gels for the separation of higher and lower molecular weight DNA fragments, respectively. φX174 RFI (5386 bp) and its nicked form φX174 RFII were purchased from New England Biolabs.

    Article Title: Genetically manipulated phages with improved pH resistance for oral administration in veterinary medicine
    Article Snippet: .. Sizes were confirmed in a 1% SGTB agarose gel using the Low Molecular Weight DNA ladder. .. Phage purification by Cesium Chloride density gradient centrifugation The mutant phages were purified by cesium chloride density gradient centrifugation.

    Article Title: Differentiated Human Midbrain-Derived Neural Progenitor Cells Express Excitatory Strychnine-Sensitive Glycine Receptors Containing ?2? Subunits
    Article Snippet: .. The correct amplicon size was asserted by agarose gel electrophoresis using low molecular weight DNA ladder (New England Biolabs, Ipswich, MA, USA). .. Oligonucleotide primers for human glycine receptor subunits α1–4 and β ( ) were designed to flank intron sequences, if feasible, using Primer 3 software.

    Article Title: High-throughput haplotype determination over long distances by Haplotype Fusion PCR and Ligation Haplotyping
    Article Snippet: .. Loading dye (Qiagen cat. no. 239901) Low molecular weight DNA ladder (NEB cat. no. N3233L) 8μl formamide (Sigma cat. no. F5786) – CAUTION: toxic – handle with care and avoid breathing vapour. .. 1μl GeneScan ROX 350 size standard (Applied Biosystems cat. no. 401735).

    Article Title: Blockade of Eosinophil-Induced Bronchial Epithelial-Mesenchymal Transition with a Geranyl Acetophenone in a Coculture Model
    Article Snippet: .. PCR products in each gel electrophoresis were ran in parallel to a Low Molecular Weight DNA Ladder (NEB, USA). .. Band intensities were quantified by ImageJ Image Processing Software (NIH, USA) and normalized by comparison to the RT-PCR product of β-actin mRNA.

    Article Title: ConcatSeq: A method for increasing throughput of single molecule sequencing by concatenating short DNA fragments
    Article Snippet: .. Low molecular weight DNA ladder was purchased from New England BioLabs (N3233). .. Oligonucleotides and Nuclease-Free Duplex Buffer were purchased from Integrated DNA Technologies.

    Nucleic Acid Electrophoresis:

    Article Title: Dual coding potential of a 2′,5′-branched ribonucleotide in DNA
    Article Snippet: .. Single-stranded oligonucleotides synthesized by Eurofins MWG Operon or a Low Molecular Weight DNA Ladder (New England BioLabs, NEB) were used as size markers in gel electrophoresis. ..

    Article Title: Blockade of Eosinophil-Induced Bronchial Epithelial-Mesenchymal Transition with a Geranyl Acetophenone in a Coculture Model
    Article Snippet: .. PCR products in each gel electrophoresis were ran in parallel to a Low Molecular Weight DNA Ladder (NEB, USA). .. Band intensities were quantified by ImageJ Image Processing Software (NIH, USA) and normalized by comparison to the RT-PCR product of β-actin mRNA.

    Isolation:

    Article Title: A LysR-family transcriptional regulator required for virulence in Brucella abortus is highly conserved among the α-proteobacteria
    Article Snippet: RNA was isolated from Brucella cultures as described previously ( ). .. A low-molecular-weight DNA ladder (New England BioLabs, Ipswich, MA, USA) was labeled with [γ-32 P]-ATP (Perkin-Elmer, San Jose, CA, USA) and polynucleotide kinase.

    Article Title: Sibling sRNA RyfA1 Influences Shigella dysenteriae Pathogenesis
    Article Snippet: Northern Blot Following growth at 37 °C in a shaking incubator, RNA was isolated as described above. .. A Low Molecular Weight DNA Ladder (New England BioLabs, Inc.) was labeled with [ɣ-32P]-ATP using polynucleotide kinase and run on the same gel as the RNA sample.

    Article Title: Differentiated Human Midbrain-Derived Neural Progenitor Cells Express Excitatory Strychnine-Sensitive Glycine Receptors Containing ?2? Subunits
    Article Snippet: Quantitative Real-time PCR After 1 and 3 weeks of differentiation in vitro , human mesencephalic NPCs from 3 cell lines were harvested and total RNA isolated using the Trizol reagent (Invitrogen). .. The correct amplicon size was asserted by agarose gel electrophoresis using low molecular weight DNA ladder (New England Biolabs, Ipswich, MA, USA).

    Article Title: Endoribonuclease YbeY Is Linked to Proper Cellular Morphology and Virulence in Brucella abortus
    Article Snippet: RNA was isolated from Brucella cultures as described previously ( ). .. A low-molecular-weight DNA ladder (New England BioLabs, Ipswich, MA) was labeled with [γ-32 P]ATP and polynucleotide kinase, and this radiolabeled ladder was also separated on the polyacrylamide gel.

    Size-exclusion Chromatography:

    Article Title: Differentiated Human Midbrain-Derived Neural Progenitor Cells Express Excitatory Strychnine-Sensitive Glycine Receptors Containing ?2? Subunits
    Article Snippet: Quantitative real-time PCR was done using cDNA from 30 ng total RNA, 0.6 µM forward and reverse primers, Platinum-SYBR Green® qPCR Supermix (Invitrogen), and 100 nM 6-carboxy-X-rhodamine (ROX) using the following protocol in an MX 3000P instrument (Stratagene, La Jolla, CA, USA): 2 min 50°C, 2 min 95°C and 50 cycles of 15 sec 95°C, 30 sec 60°C. .. The correct amplicon size was asserted by agarose gel electrophoresis using low molecular weight DNA ladder (New England Biolabs, Ipswich, MA, USA).

    Labeling:

    Article Title: A LysR-family transcriptional regulator required for virulence in Brucella abortus is highly conserved among the α-proteobacteria
    Article Snippet: .. A low-molecular-weight DNA ladder (New England BioLabs, Ipswich, MA, USA) was labeled with [γ-32 P]-ATP (Perkin-Elmer, San Jose, CA, USA) and polynucleotide kinase. ..

    Article Title: Sibling sRNA RyfA1 Influences Shigella dysenteriae Pathogenesis
    Article Snippet: .. A Low Molecular Weight DNA Ladder (New England BioLabs, Inc.) was labeled with [ɣ-32P]-ATP using polynucleotide kinase and run on the same gel as the RNA sample. ..

    Article Title: Endoribonuclease YbeY Is Linked to Proper Cellular Morphology and Virulence in Brucella abortus
    Article Snippet: .. A low-molecular-weight DNA ladder (New England BioLabs, Ipswich, MA) was labeled with [γ-32 P]ATP and polynucleotide kinase, and this radiolabeled ladder was also separated on the polyacrylamide gel. .. Following electrophoresis in 1× TBE buffer, the ladder and RNA samples were transferred to an Amersham Hybond-N+ membrane (GE Healthcare, Piscataway, NJ) in 1× TBE buffer.

    Polymerase Chain Reaction:

    Article Title: Experimental discovery of sRNAs in Vibrio cholerae by direct cloning, 5S/tRNA depletion and parallel sequencing
    Article Snippet: Generally, northern blots were performed with RNA probes transcribed from PCR-derived templates ( Supplementary Table 4 ) with T7 promoters using 32 P-UTP and T7 polymerase (Promega) according to the manufacturer's instructions. .. Low molecular weight DNA ladder (NEB) was run alongside RNA samples to provide estimations for the sizes of RNA bands.

    Article Title: Dual coding potential of a 2′,5′-branched ribonucleotide in DNA
    Article Snippet: Five microliters of the PCR reaction was mixed with 0.5 µL 10× loading-dye (25% Ficoll-400 and 0.4% xylene cyanol) and electrophoresed on a 4% agarose gel in 1× TAE buffer (40 mM Tris-acetate and 1 mM EDTA, pH 8.3). .. Single-stranded oligonucleotides synthesized by Eurofins MWG Operon or a Low Molecular Weight DNA Ladder (New England BioLabs, NEB) were used as size markers in gel electrophoresis.

    Article Title: Differentiated Human Midbrain-Derived Neural Progenitor Cells Express Excitatory Strychnine-Sensitive Glycine Receptors Containing ?2? Subunits
    Article Snippet: The correct amplicon size was asserted by agarose gel electrophoresis using low molecular weight DNA ladder (New England Biolabs, Ipswich, MA, USA). .. Cycle threshold (Ct) values were placed within the exponential phase of the PCR as described previously .

    Article Title: Blockade of Eosinophil-Induced Bronchial Epithelial-Mesenchymal Transition with a Geranyl Acetophenone in a Coculture Model
    Article Snippet: .. PCR products in each gel electrophoresis were ran in parallel to a Low Molecular Weight DNA Ladder (NEB, USA). .. Band intensities were quantified by ImageJ Image Processing Software (NIH, USA) and normalized by comparison to the RT-PCR product of β-actin mRNA.

    Plasmid Preparation:

    Article Title: RNase H1 directs origin-specific initiation of DNA replication in human mitochondria
    Article Snippet: All transcription reaction volumes were 25 μL and contained 25 mM Tris-HCl pH 8.0, 10 mM MgCl2 , 64 mM NaCl, 100 μg/mL BSA, 10 mM DTT, 400 μM ATP, 150 μM GTP, 150 μM CTP 10 μM UTP, 0.027 μM α-[32 P]UTP (3000 Ci/mmol), 4 nM of indicated plasmid template, 20 nM POLRMT, 200 nM TFAM, 60 nM TFB2M, and 40 nM TEFM where indicated. .. Low Molecular Weight DNA Ladder (NEB) was used as a size marker.

    Software:

    Article Title: Differentiated Human Midbrain-Derived Neural Progenitor Cells Express Excitatory Strychnine-Sensitive Glycine Receptors Containing ?2? Subunits
    Article Snippet: The correct amplicon size was asserted by agarose gel electrophoresis using low molecular weight DNA ladder (New England Biolabs, Ipswich, MA, USA). .. Oligonucleotide primers for human glycine receptor subunits α1–4 and β ( ) were designed to flank intron sequences, if feasible, using Primer 3 software.

    Article Title: Blockade of Eosinophil-Induced Bronchial Epithelial-Mesenchymal Transition with a Geranyl Acetophenone in a Coculture Model
    Article Snippet: PCR products in each gel electrophoresis were ran in parallel to a Low Molecular Weight DNA Ladder (NEB, USA). .. Band intensities were quantified by ImageJ Image Processing Software (NIH, USA) and normalized by comparison to the RT-PCR product of β-actin mRNA.

    Real-time Polymerase Chain Reaction:

    Article Title: Differentiated Human Midbrain-Derived Neural Progenitor Cells Express Excitatory Strychnine-Sensitive Glycine Receptors Containing ?2? Subunits
    Article Snippet: Paragraph title: Quantitative Real-time PCR ... The correct amplicon size was asserted by agarose gel electrophoresis using low molecular weight DNA ladder (New England Biolabs, Ipswich, MA, USA).

    Agarose Gel Electrophoresis:

    Article Title: Dual coding potential of a 2′,5′-branched ribonucleotide in DNA
    Article Snippet: Five microliters of the PCR reaction was mixed with 0.5 µL 10× loading-dye (25% Ficoll-400 and 0.4% xylene cyanol) and electrophoresed on a 4% agarose gel in 1× TAE buffer (40 mM Tris-acetate and 1 mM EDTA, pH 8.3). .. Single-stranded oligonucleotides synthesized by Eurofins MWG Operon or a Low Molecular Weight DNA Ladder (New England BioLabs, NEB) were used as size markers in gel electrophoresis.

    Article Title: Genetically manipulated phages with improved pH resistance for oral administration in veterinary medicine
    Article Snippet: .. Sizes were confirmed in a 1% SGTB agarose gel using the Low Molecular Weight DNA ladder. .. Phage purification by Cesium Chloride density gradient centrifugation The mutant phages were purified by cesium chloride density gradient centrifugation.

    Article Title: Differentiated Human Midbrain-Derived Neural Progenitor Cells Express Excitatory Strychnine-Sensitive Glycine Receptors Containing ?2? Subunits
    Article Snippet: .. The correct amplicon size was asserted by agarose gel electrophoresis using low molecular weight DNA ladder (New England Biolabs, Ipswich, MA, USA). .. Oligonucleotide primers for human glycine receptor subunits α1–4 and β ( ) were designed to flank intron sequences, if feasible, using Primer 3 software.

    Article Title: Blockade of Eosinophil-Induced Bronchial Epithelial-Mesenchymal Transition with a Geranyl Acetophenone in a Coculture Model
    Article Snippet: The reaction products from PCR were examined by 1.8% agarose gel electrophoresis containing 0.01% of ethidium bromide. .. PCR products in each gel electrophoresis were ran in parallel to a Low Molecular Weight DNA Ladder (NEB, USA).

    In Vitro:

    Article Title: RNase H1 directs origin-specific initiation of DNA replication in human mitochondria
    Article Snippet: Paragraph title: In vitro transcription ... Low Molecular Weight DNA Ladder (NEB) was used as a size marker.

    Article Title: Differentiated Human Midbrain-Derived Neural Progenitor Cells Express Excitatory Strychnine-Sensitive Glycine Receptors Containing ?2? Subunits
    Article Snippet: Quantitative Real-time PCR After 1 and 3 weeks of differentiation in vitro , human mesencephalic NPCs from 3 cell lines were harvested and total RNA isolated using the Trizol reagent (Invitrogen). .. The correct amplicon size was asserted by agarose gel electrophoresis using low molecular weight DNA ladder (New England Biolabs, Ipswich, MA, USA).

    Ethanol Precipitation:

    Article Title: RNase H1 directs origin-specific initiation of DNA replication in human mitochondria
    Article Snippet: The transcripts were recovered by ethanol precipitation and the pellets were dissolved in 20 μL gel loading buffer (98% formamide, 10 mM EDTA, 0.025% xylene cyanol FF, and 0.025% bromophenol blue) and heated at 95°C for 3 min. .. Low Molecular Weight DNA Ladder (NEB) was used as a size marker.

    Spectrophotometry:

    Article Title: Dual coding potential of a 2′,5′-branched ribonucleotide in DNA
    Article Snippet: DNA was quantified by UV absorbance using a Nanodrop spectrophotometer (NanoDrop 2000, Thermo Scientific). .. Single-stranded oligonucleotides synthesized by Eurofins MWG Operon or a Low Molecular Weight DNA Ladder (New England BioLabs, NEB) were used as size markers in gel electrophoresis.

    Activation Assay:

    Article Title: Blockade of Eosinophil-Induced Bronchial Epithelial-Mesenchymal Transition with a Geranyl Acetophenone in a Coculture Model
    Article Snippet: The master mix was performed in an Eppendorf thermal cycler (Eppendorf, Germany) for reverse transcription at 50°C for 30 min, initial PCR activation at 95°C for 15 min, denaturation at 94°C for 30 s, annealing at 57°C for E-cadherin, collagen I, fibronectin and β-actin for 30 s, 59°C for vimentin and β-actin for 30 s, and 65°C for TGF-β and β-actin for 30 s, elongation at 72°C for 1 min and final extension at 72°C for 10 min. .. PCR products in each gel electrophoresis were ran in parallel to a Low Molecular Weight DNA Ladder (NEB, USA).

    Marker:

    Article Title: RNase H1 directs origin-specific initiation of DNA replication in human mitochondria
    Article Snippet: .. Low Molecular Weight DNA Ladder (NEB) was used as a size marker. ..

    Staining:

    Article Title: Dual coding potential of a 2′,5′-branched ribonucleotide in DNA
    Article Snippet: Agarose gels were stained with ethidium bromide solution (0.5 µg/mL) and imaged by the Typhoon FLA 9500 (GE Healthcare). .. Single-stranded oligonucleotides synthesized by Eurofins MWG Operon or a Low Molecular Weight DNA Ladder (New England BioLabs, NEB) were used as size markers in gel electrophoresis.

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    New England Biolabs molecular weight dna ladder
    Transcription analysis of pedA . pedA -reverse transcription-PCR on cDNA from P. acidilactici bac- (1, 2, 3) or UVA1 (7, 8, 9) after 1 h 30, 2 h 30 and 3 h 30 of growth, respectively, and from P. acidilactici DSM 20284 T (4) or UL5 (6) and P. pentosaceus DSM 20336 T (5) after 2 h 30 of growth. 10: water instead of <t>DNA.</t> lm: low molecular weight DNA ladder (in bp). h: <t>Tridye</t> 100-bp DNA ladder (in bp). Expected product size: 100 bp.
    Molecular Weight Dna Ladder, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 81 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Transcription analysis of pedA . pedA -reverse transcription-PCR on cDNA from P. acidilactici bac- (1, 2, 3) or UVA1 (7, 8, 9) after 1 h 30, 2 h 30 and 3 h 30 of growth, respectively, and from P. acidilactici DSM 20284 T (4) or UL5 (6) and P. pentosaceus DSM 20336 T (5) after 2 h 30 of growth. 10: water instead of DNA. lm: low molecular weight DNA ladder (in bp). h: Tridye 100-bp DNA ladder (in bp). Expected product size: 100 bp.

    Journal: BMC Biotechnology

    Article Title: Detection of the pediocin gene pedA in strains from human faeces by real-time PCR and characterization of Pediococcus acidilactici UVA1

    doi: 10.1186/1472-6750-7-55

    Figure Lengend Snippet: Transcription analysis of pedA . pedA -reverse transcription-PCR on cDNA from P. acidilactici bac- (1, 2, 3) or UVA1 (7, 8, 9) after 1 h 30, 2 h 30 and 3 h 30 of growth, respectively, and from P. acidilactici DSM 20284 T (4) or UL5 (6) and P. pentosaceus DSM 20336 T (5) after 2 h 30 of growth. 10: water instead of DNA. lm: low molecular weight DNA ladder (in bp). h: Tridye 100-bp DNA ladder (in bp). Expected product size: 100 bp.

    Article Snippet: The low molecular weight DNA ladder and Tridye 100 bp DNA-ladder (New England BioLabs, Ipswich, MA, USA) were used as size standards.

    Techniques: Polymerase Chain Reaction, BAC Assay, Molecular Weight

    UV-crosslinking bias of different RBPs dictates the suitability of different approaches for identifying binding sites A. hnRNPA1:ssDNA interface. Crystal structure of two-RRM-containing UP1 domain of hnRNP A1 (PDB ID: 2UP1; blue) in complex with a target containing its AGGG preferred recognition motif (in this case, within single-stranded DNA, ssDNA; red). Black box. Enlarged view of the DNA:protein interface. Aromatic residues (Phe-17 and Phe-59 from RRM1) that stack with the nucleobases are shown in cyan. B. eIF4AIII:RNA interface. Crystal structure of eIF4AIII (PDB ID: 2J0S; blue) complexed with RNA (red) and AMP-PNP. Black box. Enlarged view of the RNA:protein interface. Note that the RNA bases are pointing away from the bound protein. ]. D. RIPiT and CLIP yield different types of information. Top: Two similar yet compositionally distinct hypothetical multi-subunit RNPs. RBPs (blue), non-RBPs (green) and proteins unique to each complex are shown (complex A: yellow; complex B: red). Left: RIPiT can reveal the binding sites of an intact multi-subunit RNP, and can also distinguish between footprints of two compositionally similar complexes (schematics on gray background). However, RIPiT does not conclusively define direct RBP-RNA interactions (crossed-out schematic). Right: On the contrary, while CLIP reveals no information regarding the complexes an RNA-bound RBP is part of (crossed-out schematics), it can unveil the sites of direct contact between an RBP and RNA (bottom schematic).

    Journal: Methods (San Diego, Calif.)

    Article Title: RIPiT-Seq: A high-throughput approach for footprinting RNA:protein complexes

    doi: 10.1016/j.ymeth.2013.09.013

    Figure Lengend Snippet: UV-crosslinking bias of different RBPs dictates the suitability of different approaches for identifying binding sites A. hnRNPA1:ssDNA interface. Crystal structure of two-RRM-containing UP1 domain of hnRNP A1 (PDB ID: 2UP1; blue) in complex with a target containing its AGGG preferred recognition motif (in this case, within single-stranded DNA, ssDNA; red). Black box. Enlarged view of the DNA:protein interface. Aromatic residues (Phe-17 and Phe-59 from RRM1) that stack with the nucleobases are shown in cyan. B. eIF4AIII:RNA interface. Crystal structure of eIF4AIII (PDB ID: 2J0S; blue) complexed with RNA (red) and AMP-PNP. Black box. Enlarged view of the RNA:protein interface. Note that the RNA bases are pointing away from the bound protein. ]. D. RIPiT and CLIP yield different types of information. Top: Two similar yet compositionally distinct hypothetical multi-subunit RNPs. RBPs (blue), non-RBPs (green) and proteins unique to each complex are shown (complex A: yellow; complex B: red). Left: RIPiT can reveal the binding sites of an intact multi-subunit RNP, and can also distinguish between footprints of two compositionally similar complexes (schematics on gray background). However, RIPiT does not conclusively define direct RBP-RNA interactions (crossed-out schematic). Right: On the contrary, while CLIP reveals no information regarding the complexes an RNA-bound RBP is part of (crossed-out schematics), it can unveil the sites of direct contact between an RBP and RNA (bottom schematic).

    Article Snippet: In separate, but otherwise identical reactions, also label 0.1 pmole of a synthetic RNA oligonucleotide of known length, 1 µl of low molecular weight ssDNA ladder and 1 µl of NEB 100 bp DNA ladder.

    Techniques: Binding Assay, Cross-linking Immunoprecipitation

    Biochemical analysis of proteins and RNAs from the RIPiT procedure A. Western blots showing tetracycline (Tet)-mediated induction of eIF4AIII protein with the FLAG tag at its N- or C-terminus (top and bottom panels, respectively). The Tet concentration used for induction is indicated at the top of each lane; protein identities are indicated to the right. B. Levels of proteins detected by western blots in different fractions during EJC RIPiT. The table on the top indicates the different fractions from the RIPiT procedure and the antibodies used for 1 st and 2 nd IPs. The stably expressed FLAG-tag fusion protein used in each sample is indicated directly above lane. Proteins detected by western blot are indicated to the right. C. Size distribution of EJC footprints upon RNase I titration. An autoradiogram of 26% denaturing PAGE with 5′ [ 32 P]-labeled RNA fragments from base-hydrolysis of poly U 30 oligonucleotide (lane 1) or FLAG-Magoh:eIF4AIII RIPiT (lanes 2–5). RNase I concentrations used are indicated at top of each lane; nucleotide (nt) lengths are to the left. D. Quantification of desired size RNA footprints in RIPiT elution. An autoradiogram of 20% denaturing PAGE with 5′ [ 32 P]-labeled 100 bp NEB DNA ladder (lane 1), low molecular weight ssDNA ladder (lane 2), footprints of RIPiTs indicated on top (lanes 3 and 4) and a 21 nt ssRNA oligo of known specific activity (lane 5). The signal from red rectangles is quantified in comparison to that of the 21 nt oligo to estimate the amount of RIPiT footprints in the indicated size range.

    Journal: Methods (San Diego, Calif.)

    Article Title: RIPiT-Seq: A high-throughput approach for footprinting RNA:protein complexes

    doi: 10.1016/j.ymeth.2013.09.013

    Figure Lengend Snippet: Biochemical analysis of proteins and RNAs from the RIPiT procedure A. Western blots showing tetracycline (Tet)-mediated induction of eIF4AIII protein with the FLAG tag at its N- or C-terminus (top and bottom panels, respectively). The Tet concentration used for induction is indicated at the top of each lane; protein identities are indicated to the right. B. Levels of proteins detected by western blots in different fractions during EJC RIPiT. The table on the top indicates the different fractions from the RIPiT procedure and the antibodies used for 1 st and 2 nd IPs. The stably expressed FLAG-tag fusion protein used in each sample is indicated directly above lane. Proteins detected by western blot are indicated to the right. C. Size distribution of EJC footprints upon RNase I titration. An autoradiogram of 26% denaturing PAGE with 5′ [ 32 P]-labeled RNA fragments from base-hydrolysis of poly U 30 oligonucleotide (lane 1) or FLAG-Magoh:eIF4AIII RIPiT (lanes 2–5). RNase I concentrations used are indicated at top of each lane; nucleotide (nt) lengths are to the left. D. Quantification of desired size RNA footprints in RIPiT elution. An autoradiogram of 20% denaturing PAGE with 5′ [ 32 P]-labeled 100 bp NEB DNA ladder (lane 1), low molecular weight ssDNA ladder (lane 2), footprints of RIPiTs indicated on top (lanes 3 and 4) and a 21 nt ssRNA oligo of known specific activity (lane 5). The signal from red rectangles is quantified in comparison to that of the 21 nt oligo to estimate the amount of RIPiT footprints in the indicated size range.

    Article Snippet: In separate, but otherwise identical reactions, also label 0.1 pmole of a synthetic RNA oligonucleotide of known length, 1 µl of low molecular weight ssDNA ladder and 1 µl of NEB 100 bp DNA ladder.

    Techniques: Western Blot, FLAG-tag, Concentration Assay, Stable Transfection, Titration, Polyacrylamide Gel Electrophoresis, Labeling, Molecular Weight, Activity Assay

    Principle of strand displacement switchable gRNAs. a When the RNA trigger binds the SD gRNA, the 5′ extension domain occluding handle is displaced, thereby restoring the gRNA handle. Binding of Cas12a leads to cleavage of the gRNA, which removes the 5′ extension and creates an active Cas12a-gRNA complex. The domains are labeled as follows: s—separator, h—handle, t—target, X1—toehold. The overall nomenclature follows Zhang et al. 32 : A domain is denoted by a single letter. An upper index x denotes the first x nucleotides of a domain counting from its 5′ end. A lower index x denotes all but the first x nucleotides of a domain counting from its 5′ end. A combination of upper and lower indices includes those nucleotides that are present in both subdomains. A bar above the letter marks the reverse complement of a domain. The red star marks the position where Cas12a cleaves a successfully bound gRNA. b Agarose gel showing cutting of a target DNA by handle-based SD gRNAs with two different target sequences in the absence and presence of trigger RNA (uncut: 1190 bp, cut t1: 357 bp, cut t2: 506 bp). c Denaturing PAGE showing induced gRNA processing due to trigger binding. d Agarose gel showing activation of target cutting by varying amounts of trigger RNA (uncut: 1190 bp, cut: 357 bp). e Transfer function derived from the fraction of cut target by gels as shown in ( d ) ( N = 3, t -distribution two-sided 90% confidence interval). Source data are provided as a Source Data file

    Journal: Nature Communications

    Article Title: Switching the activity of Cas12a using guide RNA strand displacement circuits

    doi: 10.1038/s41467-019-09953-w

    Figure Lengend Snippet: Principle of strand displacement switchable gRNAs. a When the RNA trigger binds the SD gRNA, the 5′ extension domain occluding handle is displaced, thereby restoring the gRNA handle. Binding of Cas12a leads to cleavage of the gRNA, which removes the 5′ extension and creates an active Cas12a-gRNA complex. The domains are labeled as follows: s—separator, h—handle, t—target, X1—toehold. The overall nomenclature follows Zhang et al. 32 : A domain is denoted by a single letter. An upper index x denotes the first x nucleotides of a domain counting from its 5′ end. A lower index x denotes all but the first x nucleotides of a domain counting from its 5′ end. A combination of upper and lower indices includes those nucleotides that are present in both subdomains. A bar above the letter marks the reverse complement of a domain. The red star marks the position where Cas12a cleaves a successfully bound gRNA. b Agarose gel showing cutting of a target DNA by handle-based SD gRNAs with two different target sequences in the absence and presence of trigger RNA (uncut: 1190 bp, cut t1: 357 bp, cut t2: 506 bp). c Denaturing PAGE showing induced gRNA processing due to trigger binding. d Agarose gel showing activation of target cutting by varying amounts of trigger RNA (uncut: 1190 bp, cut: 357 bp). e Transfer function derived from the fraction of cut target by gels as shown in ( d ) ( N = 3, t -distribution two-sided 90% confidence interval). Source data are provided as a Source Data file

    Article Snippet: For both purification methods, the purified RNA was quantified by running it on an 8 M urea denaturing TBE 12% polyacrylamide (29:1 acrylamide/bis-acrylamide) gel at 120 V for 45 min and compared with an RNA ladder of known concentration (RiboRuler Low Range RNA Ladder, ThermoFisher) to determine its concentration and with a DNA ladder (Low Molecular Weight DNA Ladder, New England Biolabs) to determine its length.

    Techniques: Binding Assay, Labeling, Agarose Gel Electrophoresis, Polyacrylamide Gel Electrophoresis, Activation Assay, Derivative Assay

    Effect of supercoiling on the structure of minicircle DNA. ( a ) Individual 336 bp minicircle topoisomers were isolated and analysed by polyacrylamide gel electrophoresis in the presence of 10 mM CaCl 2 . Mr: 100 bp DNA ladder, L: minicircle linearized by EcoRV, N: minicircle nicked by Nb.BbvCI. ( b ) Projections of cryo-ET subtomograms of hydrated 336 bp DNA minicircles of the Lk =34 topoisomer. ( c ) Commonly observed shapes were open circle, open figure-8, figure-8, racquet, handcuffs, needle, and rod, each of which are shown in orthogonal views. ( d ) Other shapes observed, especially in the more highly supercoiled topoisomers. ( e ) Shape frequency distribution plot for each topoisomer population (n=number of minicircles analysed). A weighted average for each topoisomer, approximating the average degree of compactness, is denoted by the black triangle. The weighted average was calculated by assigning each conformation a value that increased in line with compactness. Open circles were given a value of 1, open figure-8 s a value of 2, figure-8 s as a value of 3, and so on. The relative fraction of each was subsequently used to determine the average degree of compactness. Lk , Δ Lk and superhelical density (σ) for each topoisomer are shown (see Supplementary Note 1 ).

    Journal: Nature Communications

    Article Title: Structural diversity of supercoiled DNA

    doi: 10.1038/ncomms9440

    Figure Lengend Snippet: Effect of supercoiling on the structure of minicircle DNA. ( a ) Individual 336 bp minicircle topoisomers were isolated and analysed by polyacrylamide gel electrophoresis in the presence of 10 mM CaCl 2 . Mr: 100 bp DNA ladder, L: minicircle linearized by EcoRV, N: minicircle nicked by Nb.BbvCI. ( b ) Projections of cryo-ET subtomograms of hydrated 336 bp DNA minicircles of the Lk =34 topoisomer. ( c ) Commonly observed shapes were open circle, open figure-8, figure-8, racquet, handcuffs, needle, and rod, each of which are shown in orthogonal views. ( d ) Other shapes observed, especially in the more highly supercoiled topoisomers. ( e ) Shape frequency distribution plot for each topoisomer population (n=number of minicircles analysed). A weighted average for each topoisomer, approximating the average degree of compactness, is denoted by the black triangle. The weighted average was calculated by assigning each conformation a value that increased in line with compactness. Open circles were given a value of 1, open figure-8 s a value of 2, figure-8 s as a value of 3, and so on. The relative fraction of each was subsequently used to determine the average degree of compactness. Lk , Δ Lk and superhelical density (σ) for each topoisomer are shown (see Supplementary Note 1 ).

    Article Snippet: BbvCI, EcoRV, Nb.BbvCI, NdeI, Nuclease Bal-31, T4 DNA Ligase, low molecular weight DNA ladder and 100 bp DNA ladder were purchased from New England Biolabs (Ipswich, MA).

    Techniques: Isolation, Polyacrylamide Gel Electrophoresis