1 kb ladder  (New England Biolabs)


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    Name:
    1 kb DNA Ladder
    Description:
    1 kb DNA Ladder 1 000 gel lanes
    Catalog Number:
    N3232L
    Price:
    214
    Category:
    DNA Ladders
    Size:
    1 000 gel lanes
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    New England Biolabs 1 kb ladder
    1 kb DNA Ladder
    1 kb DNA Ladder 1 000 gel lanes
    https://www.bioz.com/result/1 kb ladder/product/New England Biolabs
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    1 kb ladder - by Bioz Stars, 2021-04
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    Images

    1) Product Images from "Edwardsiellosis Caused by Edwardsiella ictaluri in Laboratory Populations of Zebrafish Danio rerio"

    Article Title: Edwardsiellosis Caused by Edwardsiella ictaluri in Laboratory Populations of Zebrafish Danio rerio

    Journal: Journal of aquatic animal health

    doi: 10.1080/08997659.2013.782226

    Linearized plasmid profiles of Edwardsiella ictaluri isolates. Plasmid DNA from E. ictaluri isolated from Channel Catfish or Zebrafish was digested with EcoRI or BstZ17I, respectively, and separated by 0.6% agarose gel electrophoresis using a 1-kb DNA
    Figure Legend Snippet: Linearized plasmid profiles of Edwardsiella ictaluri isolates. Plasmid DNA from E. ictaluri isolated from Channel Catfish or Zebrafish was digested with EcoRI or BstZ17I, respectively, and separated by 0.6% agarose gel electrophoresis using a 1-kb DNA

    Techniques Used: Plasmid Preparation, Isolation, Agarose Gel Electrophoresis

    2) Product Images from "PCR-based landmark unique gene (PLUG) markers effectively assign homoeologous wheat genes to A, B and D genomes"

    Article Title: PCR-based landmark unique gene (PLUG) markers effectively assign homoeologous wheat genes to A, B and D genomes

    Journal: BMC Genomics

    doi: 10.1186/1471-2164-8-135

    1% agarose gel electrophoresis of PCR products . PCR products derived from 24 PLUG primer sets were separated using a 1% agarose gel in TAE buffer. Lane numbers correspond to marker numbers indicated in Table 1. M: 2-Log DNA Ladder (New England BioLabs Inc., Ipswich, MA, USA).
    Figure Legend Snippet: 1% agarose gel electrophoresis of PCR products . PCR products derived from 24 PLUG primer sets were separated using a 1% agarose gel in TAE buffer. Lane numbers correspond to marker numbers indicated in Table 1. M: 2-Log DNA Ladder (New England BioLabs Inc., Ipswich, MA, USA).

    Techniques Used: Agarose Gel Electrophoresis, Polymerase Chain Reaction, Derivative Assay, Marker

    3) Product Images from "Two Phosphoglucomutase Paralogs Facilitate Ionophore-Triggered Secretion of the Toxoplasma Micronemes"

    Article Title: Two Phosphoglucomutase Paralogs Facilitate Ionophore-Triggered Secretion of the Toxoplasma Micronemes

    Journal: mSphere

    doi: 10.1128/mSphere.00521-17

    Subcellular localization of PRP1 through endogenous tagging. (A) Schematic representation of generating C-terminal endogenously YFP-tagged gPRP1-YFP parasites by single homologous recombination into the RHΔ ku80 parent line. (B) PCR validation of the gPRP1-YFP genotype using the primer pair shown in panel A. Lane M contains 1-kb DNA ladder (New England Biolabs). (C) Live imaging of gPRP1-YFP parasites under intracellular and extracellular conditions as indicated. PC, phase contrast. (D) Live imaging of gPRP1-YFP parasites cotransfected with markers for the IMC (IMC1-mCherry), rhoptries (TLN1-mCherry), and micronemes (MIC8-mCherry). (E) Representative images of intracellular gPRP1-YFP parasites fixed using either 100% methanol (MetOH) or 4% paraformaldehyde (PFA) stained with anti-PRP1 (αPRP1) and anti-GFP (αGFP) antisera as indicated. Note that PFA fixation destroys the costaining of GFP and PRP1 and thus destroys the PRP1 epitope(s) recognized by the specific antiserum.
    Figure Legend Snippet: Subcellular localization of PRP1 through endogenous tagging. (A) Schematic representation of generating C-terminal endogenously YFP-tagged gPRP1-YFP parasites by single homologous recombination into the RHΔ ku80 parent line. (B) PCR validation of the gPRP1-YFP genotype using the primer pair shown in panel A. Lane M contains 1-kb DNA ladder (New England Biolabs). (C) Live imaging of gPRP1-YFP parasites under intracellular and extracellular conditions as indicated. PC, phase contrast. (D) Live imaging of gPRP1-YFP parasites cotransfected with markers for the IMC (IMC1-mCherry), rhoptries (TLN1-mCherry), and micronemes (MIC8-mCherry). (E) Representative images of intracellular gPRP1-YFP parasites fixed using either 100% methanol (MetOH) or 4% paraformaldehyde (PFA) stained with anti-PRP1 (αPRP1) and anti-GFP (αGFP) antisera as indicated. Note that PFA fixation destroys the costaining of GFP and PRP1 and thus destroys the PRP1 epitope(s) recognized by the specific antiserum.

    Techniques Used: Homologous Recombination, Polymerase Chain Reaction, Imaging, Staining

    4) Product Images from "In Situ Transfection by Controlled Release of Lipoplexes using Acoustic Droplet Vaporization"

    Article Title: In Situ Transfection by Controlled Release of Lipoplexes using Acoustic Droplet Vaporization

    Journal: Advanced healthcare materials

    doi: 10.1002/adhm.201600008

    Gel electrophoresis confirming stability of plasmid after ultrasound exposures. Lane 1) pDNA; lane 2) sonicated pDNA; lane 3) lipoplex; lane 4) sonicated lipoplex; lane 5) emulsified lipoplex released by ultrasound (3.5 MHz, mechanical index (MI) = 2.5, 10 Hz pulse repetition frequency (PRF), 30 cycles); lane 6) 1 kb linear DNA ladder marker. The mass of pDNA was equivalent in all lanes.
    Figure Legend Snippet: Gel electrophoresis confirming stability of plasmid after ultrasound exposures. Lane 1) pDNA; lane 2) sonicated pDNA; lane 3) lipoplex; lane 4) sonicated lipoplex; lane 5) emulsified lipoplex released by ultrasound (3.5 MHz, mechanical index (MI) = 2.5, 10 Hz pulse repetition frequency (PRF), 30 cycles); lane 6) 1 kb linear DNA ladder marker. The mass of pDNA was equivalent in all lanes.

    Techniques Used: Nucleic Acid Electrophoresis, Plasmid Preparation, Sonication, Marker

    5) Product Images from "Functional and Structural Characterization of Novel Type of Linker Connecting Capsid and Nucleocapsid Protein Domains in Murine Leukemia Virus *"

    Article Title: Functional and Structural Characterization of Novel Type of Linker Connecting Capsid and Nucleocapsid Protein Domains in Murine Leukemia Virus *

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M116.746461

    EMSA. The indicated proteins were incubated with a 1-kb DNA ladder, and the sample aliquots were then treated with proteinase K. All the samples were analyzed by agarose gel electrophoresis. Lanes: L : 1-kb DNA ladder; 1 : the tested proteins without the
    Figure Legend Snippet: EMSA. The indicated proteins were incubated with a 1-kb DNA ladder, and the sample aliquots were then treated with proteinase K. All the samples were analyzed by agarose gel electrophoresis. Lanes: L : 1-kb DNA ladder; 1 : the tested proteins without the

    Techniques Used: Incubation, Agarose Gel Electrophoresis

    6) Product Images from "Revealing Off-Target Cleavage Specificities of Zinc Finger Nucleases by In Vitro Selection"

    Article Title: Revealing Off-Target Cleavage Specificities of Zinc Finger Nucleases by In Vitro Selection

    Journal: Nature Methods

    doi: 10.1038/nmeth.1670

    In vitro selection for ZFN-mediated cleavage Pre-selection library members are concatemers (represented by arrows) of identical ZFN target sites lacking 5′ phosphates (orange). L = left half-site; R = right half-site, S = spacer; L′, S′, R′ = complementary sequences to L, S, R. ZFN cleavage reveals a 5′ phosphate, which is required for sequencing adapter (red and blue) ligation. The only sequences that can be amplified by PCR using primers complementary to the red and blue adapters are sequences that have been cleaved twice and have adapters on both ends. DNA cleaved at adjacent sites are purified by gel electrophoresis and sequenced. A computational screening step after sequencing ensures that the filled-in spacer sequences (S and S′) are complementary and therefore from the same molecule.
    Figure Legend Snippet: In vitro selection for ZFN-mediated cleavage Pre-selection library members are concatemers (represented by arrows) of identical ZFN target sites lacking 5′ phosphates (orange). L = left half-site; R = right half-site, S = spacer; L′, S′, R′ = complementary sequences to L, S, R. ZFN cleavage reveals a 5′ phosphate, which is required for sequencing adapter (red and blue) ligation. The only sequences that can be amplified by PCR using primers complementary to the red and blue adapters are sequences that have been cleaved twice and have adapters on both ends. DNA cleaved at adjacent sites are purified by gel electrophoresis and sequenced. A computational screening step after sequencing ensures that the filled-in spacer sequences (S and S′) are complementary and therefore from the same molecule.

    Techniques Used: In Vitro, Selection, Sequencing, Ligation, Amplification, Polymerase Chain Reaction, Purification, Nucleic Acid Electrophoresis

    7) Product Images from "Effect of Denture-Related Stomatitis Fluconazole Treatment on Oral Candida albicans Susceptibility Profile and Genotypic Variability"

    Article Title: Effect of Denture-Related Stomatitis Fluconazole Treatment on Oral Candida albicans Susceptibility Profile and Genotypic Variability

    Journal: The Open Dentistry Journal

    doi: 10.2174/1874210601509010046

    Genotypic variability of Candida albicans isolates from two different patients studied by MSP-PCR with the primer M13 at the time of DRS diagnosis (T0), and 6-months after diagnosis (T6m). Molecular size markers used were DNA Ladder (m1) and pBR322 DNA-BstNI digest (m2).
    Figure Legend Snippet: Genotypic variability of Candida albicans isolates from two different patients studied by MSP-PCR with the primer M13 at the time of DRS diagnosis (T0), and 6-months after diagnosis (T6m). Molecular size markers used were DNA Ladder (m1) and pBR322 DNA-BstNI digest (m2).

    Techniques Used: Polymerase Chain Reaction

    8) Product Images from "Genotyping bacterial and fungal pathogens using sequence variation in the gene for the CCA-adding enzyme"

    Article Title: Genotyping bacterial and fungal pathogens using sequence variation in the gene for the CCA-adding enzyme

    Journal: BMC Microbiology

    doi: 10.1186/s12866-016-0670-2

    Genome-specific amplification of loop-encoding DNA sequences. a . A Vibrio -specific forward primer (fV) was used in combination with species-specific reverse primers rValg, rVpar and rVvul to selectively amplify the loop-encoding regions. 1.0 ng genomic DNA of V. alginolyticus (Va), V. parahaemolyticus (Vp) and V. vulnificus (Vv) was used as template. The respective PCR products have a length of 204, 245 and 231 bp, respectively. b . The corresponding gene region from 3.0 ng genomic DNA from A. fumigatus , A. niger and A. terreus was amplified with appropriate primers (fA with rAfum, rAnig or rAter), leading to PCR products of 333 ( A. fumigatus , A. niger ) and 301 bp ( A. terreus ). Reaction products were separated on a 2 % agarose gel and stained with ethidium bromide. N, PCR negative control. M, 50 bp DNA ladder (NEB)
    Figure Legend Snippet: Genome-specific amplification of loop-encoding DNA sequences. a . A Vibrio -specific forward primer (fV) was used in combination with species-specific reverse primers rValg, rVpar and rVvul to selectively amplify the loop-encoding regions. 1.0 ng genomic DNA of V. alginolyticus (Va), V. parahaemolyticus (Vp) and V. vulnificus (Vv) was used as template. The respective PCR products have a length of 204, 245 and 231 bp, respectively. b . The corresponding gene region from 3.0 ng genomic DNA from A. fumigatus , A. niger and A. terreus was amplified with appropriate primers (fA with rAfum, rAnig or rAter), leading to PCR products of 333 ( A. fumigatus , A. niger ) and 301 bp ( A. terreus ). Reaction products were separated on a 2 % agarose gel and stained with ethidium bromide. N, PCR negative control. M, 50 bp DNA ladder (NEB)

    Techniques Used: Amplification, Polymerase Chain Reaction, Agarose Gel Electrophoresis, Staining, Negative Control

    Multiplex PCR with individual fluorescence-labeled primers for different Vibrio strains. a . Species-specific amplification of the flexible loop-encoding DNA sequence. Indicated amounts of individual genomic DNA (1: V. alginolyticus , 2: V. parahaemolyticus , 3: V. vulnificus ) were added to the primer mix. PCR products were visualized in the agarose gel by the different fluorescence of the species-specific primers. Down to 10 pg of each DNA sample were readily detected, without any cross reactivity with the other genomes. N, negative control. b . Human DNA does not interfere with the specific detection of Vibrio DNA. 0.1 ng of genomic DNA (1: V. alginolyticus , 2: V. parahaemolyticus , 3: V. vulnificus ) were mixed with a 500 to 1,000-fold excess (50 and 100 ng) of human genomic DNA in a multiplex PCR and visualized as above. Compared to the positive control (0, no human DNA added), no additional bands appeared, indicating an exclusive and highly specific amplification of Vibrio DNA only. N, negative control with 50 ng of human genomic DNA
    Figure Legend Snippet: Multiplex PCR with individual fluorescence-labeled primers for different Vibrio strains. a . Species-specific amplification of the flexible loop-encoding DNA sequence. Indicated amounts of individual genomic DNA (1: V. alginolyticus , 2: V. parahaemolyticus , 3: V. vulnificus ) were added to the primer mix. PCR products were visualized in the agarose gel by the different fluorescence of the species-specific primers. Down to 10 pg of each DNA sample were readily detected, without any cross reactivity with the other genomes. N, negative control. b . Human DNA does not interfere with the specific detection of Vibrio DNA. 0.1 ng of genomic DNA (1: V. alginolyticus , 2: V. parahaemolyticus , 3: V. vulnificus ) were mixed with a 500 to 1,000-fold excess (50 and 100 ng) of human genomic DNA in a multiplex PCR and visualized as above. Compared to the positive control (0, no human DNA added), no additional bands appeared, indicating an exclusive and highly specific amplification of Vibrio DNA only. N, negative control with 50 ng of human genomic DNA

    Techniques Used: Multiplex Assay, Polymerase Chain Reaction, Fluorescence, Labeling, Amplification, Sequencing, Agarose Gel Electrophoresis, Negative Control, Positive Control

    9) Product Images from "Modifications and optimization of manual methods for polymerase chain reaction and 16S rRNA gene sequencing quality community DNA extraction from goat rumen digesta"

    Article Title: Modifications and optimization of manual methods for polymerase chain reaction and 16S rRNA gene sequencing quality community DNA extraction from goat rumen digesta

    Journal: Veterinary World

    doi: 10.14202/vetworld.2018.990-1000

    (a) Community DNA extraction with Enzymatic method (EM)1, Enzymatic-Chemical method (ECM)1 and Enzymatic+Chemical+Physical method (ECPM2) methods. Community DNA extraction with > 15 Kb band on 1% agarose gel electrophoresis using (b) ECPM2, EM1, and ECM1 method, Lane 1: Ladder: 1 Kb, New England Biolabs. Community DNA extraction with ECPM2 using Lane 2-5: Solid rumen digesta, Lane 6-9: Squeezed rumen digesta, EM1 using Lane 10: Solid rumen digesta, Lane 11: Blank, ECM1 using Lane 12: Solid rumen digesta, and Lane 13: Squeezed rumen digesta. (b) Community DNA extraction with CM4 method. Community DNA extraction with > 15 Kb band on 1% agarose gel electrophoresis using (a) CM4 method, Lane 1: Ladder: 1 Kb, New England Biolabs, Lane 2: Blank, Lane 3-6: Community DNA with CM4 method, Lane 7-8: Blank, Lane 9-12: Community DNA with CM4 method, and Lane 13: Blank.
    Figure Legend Snippet: (a) Community DNA extraction with Enzymatic method (EM)1, Enzymatic-Chemical method (ECM)1 and Enzymatic+Chemical+Physical method (ECPM2) methods. Community DNA extraction with > 15 Kb band on 1% agarose gel electrophoresis using (b) ECPM2, EM1, and ECM1 method, Lane 1: Ladder: 1 Kb, New England Biolabs. Community DNA extraction with ECPM2 using Lane 2-5: Solid rumen digesta, Lane 6-9: Squeezed rumen digesta, EM1 using Lane 10: Solid rumen digesta, Lane 11: Blank, ECM1 using Lane 12: Solid rumen digesta, and Lane 13: Squeezed rumen digesta. (b) Community DNA extraction with CM4 method. Community DNA extraction with > 15 Kb band on 1% agarose gel electrophoresis using (a) CM4 method, Lane 1: Ladder: 1 Kb, New England Biolabs, Lane 2: Blank, Lane 3-6: Community DNA with CM4 method, Lane 7-8: Blank, Lane 9-12: Community DNA with CM4 method, and Lane 13: Blank.

    Techniques Used: DNA Extraction, Agarose Gel Electrophoresis

    (a and b) Standard polymerase chain reaction (PCR) using community DNA extracted using modified methods. Standard PCR using (a) universal bacterial primers with community DNA extracted using Lane 1: 1 Kb ladder (New England Biolabs), Lane 2: Enzymatic method (EM)1, Lane 3: EC1, Lane 4: Enzymatic+Chemical+Physical method (ECPM)2, Lane 5: Chemical method (CM)1, Lane 6: CM4, Lane 7: CM2 methods, Lane 9: Genomic DNA of Bacillus subtilis as a positive control and Lane 10: blank (b) Specific targeted bacterial 16S rRNA gene primers and community DNA as follows. Lane 1: 100 bp ladder (New England Biolabs), Lane 2: Blank, Lane 3-4: CM4 and ECPM2 DNA with genus Bacteroides and Prevotella (418 bp), Lane 5-6: CM4 and ECPM2 DNA with Streptococcus bovis (869 bp), Lane 7-8: CM4 and ECPM2 DNA with Ruminococcus flavefaciens (835 bp), Lane 9-10: CM4 and ECPM2 DNA with Fibrobacter succinogenes (446 bp), and Lane 11-12: CM4 and ECPM2 DNA with Selenomonas ruminantium (513 bp), and Lane 13: blank.
    Figure Legend Snippet: (a and b) Standard polymerase chain reaction (PCR) using community DNA extracted using modified methods. Standard PCR using (a) universal bacterial primers with community DNA extracted using Lane 1: 1 Kb ladder (New England Biolabs), Lane 2: Enzymatic method (EM)1, Lane 3: EC1, Lane 4: Enzymatic+Chemical+Physical method (ECPM)2, Lane 5: Chemical method (CM)1, Lane 6: CM4, Lane 7: CM2 methods, Lane 9: Genomic DNA of Bacillus subtilis as a positive control and Lane 10: blank (b) Specific targeted bacterial 16S rRNA gene primers and community DNA as follows. Lane 1: 100 bp ladder (New England Biolabs), Lane 2: Blank, Lane 3-4: CM4 and ECPM2 DNA with genus Bacteroides and Prevotella (418 bp), Lane 5-6: CM4 and ECPM2 DNA with Streptococcus bovis (869 bp), Lane 7-8: CM4 and ECPM2 DNA with Ruminococcus flavefaciens (835 bp), Lane 9-10: CM4 and ECPM2 DNA with Fibrobacter succinogenes (446 bp), and Lane 11-12: CM4 and ECPM2 DNA with Selenomonas ruminantium (513 bp), and Lane 13: blank.

    Techniques Used: Polymerase Chain Reaction, Modification, Positive Control

    10) Product Images from "Targeted mutagenesis in a human-parasitic nematode"

    Article Title: Targeted mutagenesis in a human-parasitic nematode

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1006675

    CRISPR-mediated homology-directed repair of Ss-unc-22 . ( A ) Strategy for HDR at Ss-unc-22 target site #2. unc F 1 iL3s that displayed both the nicotine-twitching phenotype and red fluorescence were selected as candidates for HDR and were genotyped using the primer sets indicated. 5’ and 3’ integration primer pairs amplify only following successful integration of Ss-act-2 :: mRFPmars into site #2. HA = homology arm. ( B ) Representative DIC + epifluorescence overlays of unc F 1 iL3s expressing Ss-act-2 :: mRFPmars . Top, iL3 expressing mRFPmars (sparse expression indicated by the arrow) from an extrachromosomal array. Bottom, iL3 expressing mRFPmars following HDR, showing near-uniform mRFPmars expression in the body wall. For both images, anterior is to the left. Scale bar = 50 μm. ( C ) Representative genotypes of a wild-type iL3 and unc F 1 iL3s expressing mRFPmars . Genomic DNA from individual iL3s was split into four reactions: ctrl. = control reaction amplifying 416 bp of the first exon of the Ss-act-2 gene to confirm the presence of genomic DNA; wt = reaction for the wild-type locus of site #2 where primer R1 overlaps the predicted CRISPR cut site; 5’ = reaction for insertion of the 5’ border of the integrated cassette; 3’ = reaction for insertion of the 3’ border of the integrated cassette. For genotypes: array = red unc F 1 iL3s that showed no evidence of integration; integrated = red unc F 1 iL3s with successful HDR. Some integrated iL3s had putative homozygous disruptions of Ss-unc-22 site #2 ( e . g . iL3s #4 and #7, which lacked the wt band). Asterisks indicate genotypes for iL3s shown in B . Size markers = 2 kb, 1.5 kb, 1 kb, and 500 bp from top to bottom.
    Figure Legend Snippet: CRISPR-mediated homology-directed repair of Ss-unc-22 . ( A ) Strategy for HDR at Ss-unc-22 target site #2. unc F 1 iL3s that displayed both the nicotine-twitching phenotype and red fluorescence were selected as candidates for HDR and were genotyped using the primer sets indicated. 5’ and 3’ integration primer pairs amplify only following successful integration of Ss-act-2 :: mRFPmars into site #2. HA = homology arm. ( B ) Representative DIC + epifluorescence overlays of unc F 1 iL3s expressing Ss-act-2 :: mRFPmars . Top, iL3 expressing mRFPmars (sparse expression indicated by the arrow) from an extrachromosomal array. Bottom, iL3 expressing mRFPmars following HDR, showing near-uniform mRFPmars expression in the body wall. For both images, anterior is to the left. Scale bar = 50 μm. ( C ) Representative genotypes of a wild-type iL3 and unc F 1 iL3s expressing mRFPmars . Genomic DNA from individual iL3s was split into four reactions: ctrl. = control reaction amplifying 416 bp of the first exon of the Ss-act-2 gene to confirm the presence of genomic DNA; wt = reaction for the wild-type locus of site #2 where primer R1 overlaps the predicted CRISPR cut site; 5’ = reaction for insertion of the 5’ border of the integrated cassette; 3’ = reaction for insertion of the 3’ border of the integrated cassette. For genotypes: array = red unc F 1 iL3s that showed no evidence of integration; integrated = red unc F 1 iL3s with successful HDR. Some integrated iL3s had putative homozygous disruptions of Ss-unc-22 site #2 ( e . g . iL3s #4 and #7, which lacked the wt band). Asterisks indicate genotypes for iL3s shown in B . Size markers = 2 kb, 1.5 kb, 1 kb, and 500 bp from top to bottom.

    Techniques Used: CRISPR, Fluorescence, Activated Clotting Time Assay, Expressing

    CRISPR-mediated mutagenesis of Ss-unc-22 results in putative deletion of the target locus. ( A ) Representative gel of wild-type iL3s (top) or unc F 1 iL3s from RNP injections at site #3 (bottom). Genomic DNA from each iL3 was split into two reactions: ctrl. = control reaction amplifying 416 bp of the first exon of the Ss-act-2 gene to confirm the presence of genomic DNA; u22 = reaction amplifying 660 bp around site #3. Size markers = 1.5 kb, 1 kb, and 500 bp from top to bottom. ( B ) The Ss-unc-22 region is significantly depleted in unc F 1 iL3s. Left: relative quantity analysis of PCR products. All control bands and all u22 bands were quantified relative to their respective reference bands, denoted by asterisks in A . Values > 1 indicate more PCR product than the reference while values
    Figure Legend Snippet: CRISPR-mediated mutagenesis of Ss-unc-22 results in putative deletion of the target locus. ( A ) Representative gel of wild-type iL3s (top) or unc F 1 iL3s from RNP injections at site #3 (bottom). Genomic DNA from each iL3 was split into two reactions: ctrl. = control reaction amplifying 416 bp of the first exon of the Ss-act-2 gene to confirm the presence of genomic DNA; u22 = reaction amplifying 660 bp around site #3. Size markers = 1.5 kb, 1 kb, and 500 bp from top to bottom. ( B ) The Ss-unc-22 region is significantly depleted in unc F 1 iL3s. Left: relative quantity analysis of PCR products. All control bands and all u22 bands were quantified relative to their respective reference bands, denoted by asterisks in A . Values > 1 indicate more PCR product than the reference while values

    Techniques Used: CRISPR, Mutagenesis, Activated Clotting Time Assay, Polymerase Chain Reaction

    11) Product Images from "Is the abundance of Faecalibacterium prausnitzii relevant to Crohn's disease?"

    Article Title: Is the abundance of Faecalibacterium prausnitzii relevant to Crohn's disease?

    Journal: Fems Microbiology Letters

    doi: 10.1111/j.1574-6968.2010.02057.x

    Relative quantity of  Faecalibacterium prausnitzii  in faecal DNA determined by PCR. Agarose gel showing bands of the  F. prausnitzii  amplicon from 22 faecal samples. The upper band of 778 bp corresponds to the M21/2 subgroup; the lower 650 bp band corresponds to the A2-165 subgroup. The same amount of template DNA was added to each PCR reaction, and so these differences indicate the relative amount of  F. prausnitzii  in each patient. Where a PCR product was present, the brightness of the band was measured using the software  quantity one  (Bio-Rad) and converted to DNA amount by reference to a 1 kb ladder (New England Biolabs: right-hand lane).
    Figure Legend Snippet: Relative quantity of Faecalibacterium prausnitzii in faecal DNA determined by PCR. Agarose gel showing bands of the F. prausnitzii amplicon from 22 faecal samples. The upper band of 778 bp corresponds to the M21/2 subgroup; the lower 650 bp band corresponds to the A2-165 subgroup. The same amount of template DNA was added to each PCR reaction, and so these differences indicate the relative amount of F. prausnitzii in each patient. Where a PCR product was present, the brightness of the band was measured using the software quantity one (Bio-Rad) and converted to DNA amount by reference to a 1 kb ladder (New England Biolabs: right-hand lane).

    Techniques Used: Polymerase Chain Reaction, Agarose Gel Electrophoresis, Amplification, Software

    12) Product Images from "Sizing femtogram amounts of dsDNA by single-molecule counting"

    Article Title: Sizing femtogram amounts of dsDNA by single-molecule counting

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkv904

    Single-molecule intensity distribution of a 1 kbp DNA ladder. ( A ) intensity distribution histogram of detected molecules (red), fitted Gaussians sum (black) and the residue of the fitting (blue). ( B ) a plot of expected DNA lengths in the ladder sample versus the centers of the fitted Gaussian peaks exhibits linear dependency with R 2 = 0.99932.
    Figure Legend Snippet: Single-molecule intensity distribution of a 1 kbp DNA ladder. ( A ) intensity distribution histogram of detected molecules (red), fitted Gaussians sum (black) and the residue of the fitting (blue). ( B ) a plot of expected DNA lengths in the ladder sample versus the centers of the fitted Gaussian peaks exhibits linear dependency with R 2 = 0.99932.

    Techniques Used:

    13) Product Images from "Ability of Polyphosphate and Nucleic Acids to Trigger Blood Clotting: Some Observations and Caveats"

    Article Title: Ability of Polyphosphate and Nucleic Acids to Trigger Blood Clotting: Some Observations and Caveats

    Journal: Frontiers in Medicine

    doi: 10.3389/fmed.2018.00107

    Traces of polyphosphate (polyP) in cell-derived DNA. DNA isolated from HEK 293 cells using the DNeasy Blood Tissue kit was extensively digested with benzonase to hydrolyze the DNA, concentrated, and then resolved by electrophoresis on a 4–20% polyacrylamide gel. Samples were: a locally prepared polyP ladder (lengths indicated in phosphate units); DNA purified from HEK 293 cells and digested with benzonase (“Pre”); the same material following digestion with calf intestinal alkaline phosphatase (CIAP); and 50 bp DNA ladder. The same gel was stained sequentially, using: (A) DAPI with extended photobleaching to detect polyP ( 27 ) and (B) SYBR Green I to detect DNA (after removing DAPI by repeated rinsing). The material in the lane marked “Pre” is clearly polyP as it photobleached rapidly (A) , was digested by CIAP (A) , and did not stain with SYBR Green 1 (B) .
    Figure Legend Snippet: Traces of polyphosphate (polyP) in cell-derived DNA. DNA isolated from HEK 293 cells using the DNeasy Blood Tissue kit was extensively digested with benzonase to hydrolyze the DNA, concentrated, and then resolved by electrophoresis on a 4–20% polyacrylamide gel. Samples were: a locally prepared polyP ladder (lengths indicated in phosphate units); DNA purified from HEK 293 cells and digested with benzonase (“Pre”); the same material following digestion with calf intestinal alkaline phosphatase (CIAP); and 50 bp DNA ladder. The same gel was stained sequentially, using: (A) DAPI with extended photobleaching to detect polyP ( 27 ) and (B) SYBR Green I to detect DNA (after removing DAPI by repeated rinsing). The material in the lane marked “Pre” is clearly polyP as it photobleached rapidly (A) , was digested by CIAP (A) , and did not stain with SYBR Green 1 (B) .

    Techniques Used: Derivative Assay, Isolation, Electrophoresis, Purification, Staining, SYBR Green Assay

    14) Product Images from "Contamination sources, serogroups, biofilm-forming ability and biocide resistance of Listeria monocytogenes persistent in tilapia-processing facilities"

    Article Title: Contamination sources, serogroups, biofilm-forming ability and biocide resistance of Listeria monocytogenes persistent in tilapia-processing facilities

    Journal: Journal of Food Science and Technology

    doi: 10.1007/s13197-017-2843-x

    Agarose gels showing RAPD patterns of L. monocytogenes for primers DAF4 ( a ), HLWL85 ( b ) and OPM-01 ( c ). Lane 1: DNA molecular weight marker (100 bp DNA ladder, 100–1517 bp, New England Biolabs)
    Figure Legend Snippet: Agarose gels showing RAPD patterns of L. monocytogenes for primers DAF4 ( a ), HLWL85 ( b ) and OPM-01 ( c ). Lane 1: DNA molecular weight marker (100 bp DNA ladder, 100–1517 bp, New England Biolabs)

    Techniques Used: Molecular Weight, Marker

    15) Product Images from "Two Phosphoglucomutase Paralogs Facilitate Ionophore-Triggered Secretion of the Toxoplasma Micronemes"

    Article Title: Two Phosphoglucomutase Paralogs Facilitate Ionophore-Triggered Secretion of the Toxoplasma Micronemes

    Journal: mSphere

    doi: 10.1128/mSphere.00521-17

    Subcellular localization of PRP1 through endogenous tagging. (A) Schematic representation of generating C-terminal endogenously YFP-tagged gPRP1-YFP parasites by single homologous recombination into the RHΔ ku80 parent line. (B) PCR validation of the gPRP1-YFP genotype using the primer pair shown in panel A. Lane M contains 1-kb DNA ladder (New England Biolabs). (C) Live imaging of gPRP1-YFP parasites under intracellular and extracellular conditions as indicated. PC, phase contrast. (D) Live imaging of gPRP1-YFP parasites cotransfected with markers for the IMC (IMC1-mCherry), rhoptries (TLN1-mCherry), and micronemes (MIC8-mCherry). (E) Representative images of intracellular gPRP1-YFP parasites fixed using either 100% methanol (MetOH) or 4% paraformaldehyde (PFA) stained with anti-PRP1 (αPRP1) and anti-GFP (αGFP) antisera as indicated. Note that PFA fixation destroys the costaining of GFP and PRP1 and thus destroys the PRP1 epitope(s) recognized by the specific antiserum.
    Figure Legend Snippet: Subcellular localization of PRP1 through endogenous tagging. (A) Schematic representation of generating C-terminal endogenously YFP-tagged gPRP1-YFP parasites by single homologous recombination into the RHΔ ku80 parent line. (B) PCR validation of the gPRP1-YFP genotype using the primer pair shown in panel A. Lane M contains 1-kb DNA ladder (New England Biolabs). (C) Live imaging of gPRP1-YFP parasites under intracellular and extracellular conditions as indicated. PC, phase contrast. (D) Live imaging of gPRP1-YFP parasites cotransfected with markers for the IMC (IMC1-mCherry), rhoptries (TLN1-mCherry), and micronemes (MIC8-mCherry). (E) Representative images of intracellular gPRP1-YFP parasites fixed using either 100% methanol (MetOH) or 4% paraformaldehyde (PFA) stained with anti-PRP1 (αPRP1) and anti-GFP (αGFP) antisera as indicated. Note that PFA fixation destroys the costaining of GFP and PRP1 and thus destroys the PRP1 epitope(s) recognized by the specific antiserum.

    Techniques Used: Homologous Recombination, Polymerase Chain Reaction, Imaging, Staining

    16) Product Images from "Molecular and Phylogenetic analysis revealed new genotypes of Theileria annulata parasites from India"

    Article Title: Molecular and Phylogenetic analysis revealed new genotypes of Theileria annulata parasites from India

    Journal: Parasites & Vectors

    doi: 10.1186/s13071-015-1075-z

    PCR amplification in cattle DNA samples. Agarose gel electrophoresis of amplified DNA from different cattle blood DNA samples by using 18S rRNA. Lanes: 1-Negative control distill water: Lane 2- T. orientalis positive DNA sample; Lane 3 to Lane 8 positive blood DNA samples from cattle; Lane 9–100 base pairs DNA ladder
    Figure Legend Snippet: PCR amplification in cattle DNA samples. Agarose gel electrophoresis of amplified DNA from different cattle blood DNA samples by using 18S rRNA. Lanes: 1-Negative control distill water: Lane 2- T. orientalis positive DNA sample; Lane 3 to Lane 8 positive blood DNA samples from cattle; Lane 9–100 base pairs DNA ladder

    Techniques Used: Polymerase Chain Reaction, Amplification, Agarose Gel Electrophoresis, Negative Control

    17) Product Images from "Generating Transgenic Mice from Bacterial Artificial Chromosomes: Transgenesis Efficiency, Integration and Expression Outcomes"

    Article Title: Generating Transgenic Mice from Bacterial Artificial Chromosomes: Transgenesis Efficiency, Integration and Expression Outcomes

    Journal: Transgenic research

    doi: 10.1007/s11248-009-9271-2

    Pulsed-Field Gel Analysis of BAC DNA. 1a. Analysis of intact circular BAC DNA and sheared BAC DNA. Lane 1: Midrange PFG Marker II (New England Biolabs). Lanes 2, 3, and 4: BAC 2039 (expected size 182kb). Lane 2: Circular BAC DNA: 0.34 ug. Lane 3: Circular BAC DNA: 3.4 ug. Lane 4: Not I cut BAC DNA. Lane 5: deliberately empty. Lanes 6, 7, and 8 contain three fractions from a size exclusion column for BAC 2053. The expected size fragment is 140 kb. All three fractions contain sheared DNA smaller than 97 kb instead of intact linearized BAC DNA. 1b. Comparative analysis of sheared BAC DNA with conventional gel electrophoresis and pulsed-field gel electrophoresis. Lane 1: 2-Log DNA Ladder (New England Biolabs) separation on a conventional 0.8% agarose gel. Lane 2. Sheared BAC DNA runs as a sharp band on a conventional gel. Lane 3. Midrange PFG Marker II (New England Biolabs) separation on a pulsed-field gel. Lane 4. The same sheared BAC DNA that resolves as a sharp band on a conventional gel (Lane 2) appears as a smear upon pulsed-field gel electrophoresis. Arrow indicates expected size of BAC in Lane 4.
    Figure Legend Snippet: Pulsed-Field Gel Analysis of BAC DNA. 1a. Analysis of intact circular BAC DNA and sheared BAC DNA. Lane 1: Midrange PFG Marker II (New England Biolabs). Lanes 2, 3, and 4: BAC 2039 (expected size 182kb). Lane 2: Circular BAC DNA: 0.34 ug. Lane 3: Circular BAC DNA: 3.4 ug. Lane 4: Not I cut BAC DNA. Lane 5: deliberately empty. Lanes 6, 7, and 8 contain three fractions from a size exclusion column for BAC 2053. The expected size fragment is 140 kb. All three fractions contain sheared DNA smaller than 97 kb instead of intact linearized BAC DNA. 1b. Comparative analysis of sheared BAC DNA with conventional gel electrophoresis and pulsed-field gel electrophoresis. Lane 1: 2-Log DNA Ladder (New England Biolabs) separation on a conventional 0.8% agarose gel. Lane 2. Sheared BAC DNA runs as a sharp band on a conventional gel. Lane 3. Midrange PFG Marker II (New England Biolabs) separation on a pulsed-field gel. Lane 4. The same sheared BAC DNA that resolves as a sharp band on a conventional gel (Lane 2) appears as a smear upon pulsed-field gel electrophoresis. Arrow indicates expected size of BAC in Lane 4.

    Techniques Used: Pulsed-Field Gel, BAC Assay, Marker, Nucleic Acid Electrophoresis, Electrophoresis, Agarose Gel Electrophoresis

    18) Product Images from "Airway CD8+ T cells induced by pulmonary DNA immunization mediate protective anti-viral immunity"

    Article Title: Airway CD8+ T cells induced by pulmonary DNA immunization mediate protective anti-viral immunity

    Journal: Mucosal Immunology

    doi: 10.1038/mi.2012.59

    Persistence of pulmonary CD8 + T cells is independent of the duration of antigen presentation in the airways and lungs or local T-cell proliferation. ( a ) Lungs were harvested from B6 mice 2 days and 3 weeks post pulmonary PEI-DNA-OVA immunization and plasmid DNA isolated from the tissue. The OVA sequence was amplified from plasmid DNA by PCR. The plasmid DNA-OVA used for immunization served as positive control (pos ctr) while plasmid DNA without the OVA insert served as the negative control (neg ctr). ( b ) B6.PL (Thy 1.1 + ) mice were immunized by the pulmonary route with PEI-DNA-OVA and 21 and 42 days later, received adoptively transferred labeled OT-I cells (Thy 1.2 + ) by intravenous injection. The dilution of eFluor 670 in donor Thy1.2 + tetramer + CD8 + T cells was evaluated in mediastinal lymph nodes (MLN) of the immunized mice 4 days post transfer. Representative histograms show the average percentages±s.e. of proliferating Thy1.2 + tetramer + CD8 + T cells (3–4 mice per group). ( c ) Labeled OT-I cells (Thy 1.2 + ) were adoptively transferred to the airway lumen of B6.PL mice (Thy 1.1 + ). One day later, mice were immunized by the pulmonary route with PEI-DNA-OVA, and 10 days later, donor tetramer-binding CD8 + T cells were isolated from the BAL and lungs, and evaluated for the dilution of eFluor 670 staining. ( d ) B6-PL (Thy 1.1 + ) mice were immunized by the pulmonary route with PEI-DNA-OVA and 21 days later received adoptively transferred labeled OT-I cells (Thy 1.2 + ) in their airway lumens. The dilution of eFluor 670 in donor Thy1.2 + tetramer + CD8 + T cells was evaluated 4 days later in BAL and lungs. Representative histograms show the average percentages±s.e. of proliferating Thy1.2 + tetramer + CD8 + T cells (6–8 mice per group). Local antigen presentation was also evaluated in lungs of B6 mice immunized by the pulmonary route with PEI-DNA-OVA. Lungs were harvested 6 weeks following immunization and co-cultured with the RF.33.70 hybridoma overnight. The levels of IL-2 secretion from the RF33.70 hybridoma in response to SIINFEKL presentation were determined using a mouse IL-2 enzyme-linked immunosorbent assay. Bars represent the average±s.e. of secreted IL-2 (3 mice per group). ( e ) Mice were inoculated by the pulmonary route with PEI-DNA-gp120, and 3 and 6 weeks later eFluor 670 was applied to the airways to stain local resident T cells. Four days later, the dilution of eFluor 670 staining of tetramer + CD8 + T cells in the BAL was assessed. Histograms representative of 3–8 mice per time point are shown. Proliferation of antigen-specific CD8 + T cells was also evaluated in the BAL of immunized Balb/c mice 6 weeks following pulmonary PEI-DNA-gp120 immunization. Mice were injected with 250 μg 5-ethynyl-2′-deoxyuridine (EdU) intraperitoneally 12 h before sacrifice. Cells isolated from the BAL were stained with monoclonal antibodies to CD4 and CD8, fixed, permeabilized, and EdU detected with the Alexa Fluor 647 Click-iT EdU flow cytometry assay kit. Representative plot show the average percentages±s.e. of EdU incorporation by BAL CD8 + T cells (3 mice per group). BAL CD8 + T cells from mice infected intranasally with rVac-gp160 were used as a positive control for local proliferation. APC, antigen-presenting cells; BAL, broncho-alveolar lavage; ctr, control; IL-2, interleukin 2; pep, peptide; PEI-DNA-OVA, polyethyleneimine-DNA-Ovalbumin complexes.
    Figure Legend Snippet: Persistence of pulmonary CD8 + T cells is independent of the duration of antigen presentation in the airways and lungs or local T-cell proliferation. ( a ) Lungs were harvested from B6 mice 2 days and 3 weeks post pulmonary PEI-DNA-OVA immunization and plasmid DNA isolated from the tissue. The OVA sequence was amplified from plasmid DNA by PCR. The plasmid DNA-OVA used for immunization served as positive control (pos ctr) while plasmid DNA without the OVA insert served as the negative control (neg ctr). ( b ) B6.PL (Thy 1.1 + ) mice were immunized by the pulmonary route with PEI-DNA-OVA and 21 and 42 days later, received adoptively transferred labeled OT-I cells (Thy 1.2 + ) by intravenous injection. The dilution of eFluor 670 in donor Thy1.2 + tetramer + CD8 + T cells was evaluated in mediastinal lymph nodes (MLN) of the immunized mice 4 days post transfer. Representative histograms show the average percentages±s.e. of proliferating Thy1.2 + tetramer + CD8 + T cells (3–4 mice per group). ( c ) Labeled OT-I cells (Thy 1.2 + ) were adoptively transferred to the airway lumen of B6.PL mice (Thy 1.1 + ). One day later, mice were immunized by the pulmonary route with PEI-DNA-OVA, and 10 days later, donor tetramer-binding CD8 + T cells were isolated from the BAL and lungs, and evaluated for the dilution of eFluor 670 staining. ( d ) B6-PL (Thy 1.1 + ) mice were immunized by the pulmonary route with PEI-DNA-OVA and 21 days later received adoptively transferred labeled OT-I cells (Thy 1.2 + ) in their airway lumens. The dilution of eFluor 670 in donor Thy1.2 + tetramer + CD8 + T cells was evaluated 4 days later in BAL and lungs. Representative histograms show the average percentages±s.e. of proliferating Thy1.2 + tetramer + CD8 + T cells (6–8 mice per group). Local antigen presentation was also evaluated in lungs of B6 mice immunized by the pulmonary route with PEI-DNA-OVA. Lungs were harvested 6 weeks following immunization and co-cultured with the RF.33.70 hybridoma overnight. The levels of IL-2 secretion from the RF33.70 hybridoma in response to SIINFEKL presentation were determined using a mouse IL-2 enzyme-linked immunosorbent assay. Bars represent the average±s.e. of secreted IL-2 (3 mice per group). ( e ) Mice were inoculated by the pulmonary route with PEI-DNA-gp120, and 3 and 6 weeks later eFluor 670 was applied to the airways to stain local resident T cells. Four days later, the dilution of eFluor 670 staining of tetramer + CD8 + T cells in the BAL was assessed. Histograms representative of 3–8 mice per time point are shown. Proliferation of antigen-specific CD8 + T cells was also evaluated in the BAL of immunized Balb/c mice 6 weeks following pulmonary PEI-DNA-gp120 immunization. Mice were injected with 250 μg 5-ethynyl-2′-deoxyuridine (EdU) intraperitoneally 12 h before sacrifice. Cells isolated from the BAL were stained with monoclonal antibodies to CD4 and CD8, fixed, permeabilized, and EdU detected with the Alexa Fluor 647 Click-iT EdU flow cytometry assay kit. Representative plot show the average percentages±s.e. of EdU incorporation by BAL CD8 + T cells (3 mice per group). BAL CD8 + T cells from mice infected intranasally with rVac-gp160 were used as a positive control for local proliferation. APC, antigen-presenting cells; BAL, broncho-alveolar lavage; ctr, control; IL-2, interleukin 2; pep, peptide; PEI-DNA-OVA, polyethyleneimine-DNA-Ovalbumin complexes.

    Techniques Used: Mouse Assay, Plasmid Preparation, Isolation, Sequencing, Amplification, Polymerase Chain Reaction, Positive Control, Negative Control, Labeling, Injection, Binding Assay, Staining, Cell Culture, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Cytometry, Infection

    19) Product Images from "Escalating Association of Vibrio cholerae O139 with Cholera Outbreaks in India"

    Article Title: Escalating Association of Vibrio cholerae O139 with Cholera Outbreaks in India

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.40.7.2635-2637.2002

    RAPD profiles of outbreak strains using primer 1281. (a) Lanes 1 and 11, 1-kb ladder; lanes 2 and 3, Orissa, 1999; lanes 4 and 5, Ahmedabad, 2000; lanes 6 and 7, Karnataka, 2000; lanes 8, 9, and 10, Hyderabad, 2000. (b) Lanes 1 and 11, 1-kb ladder; lane 2, O139 reference strain ATCC 51394 (originally designated MO45); lanes 3, 4, 5, and 6, Orissa, 2000; lanes 7, 8, 9, and 10, Calcutta, 2000.
    Figure Legend Snippet: RAPD profiles of outbreak strains using primer 1281. (a) Lanes 1 and 11, 1-kb ladder; lanes 2 and 3, Orissa, 1999; lanes 4 and 5, Ahmedabad, 2000; lanes 6 and 7, Karnataka, 2000; lanes 8, 9, and 10, Hyderabad, 2000. (b) Lanes 1 and 11, 1-kb ladder; lane 2, O139 reference strain ATCC 51394 (originally designated MO45); lanes 3, 4, 5, and 6, Orissa, 2000; lanes 7, 8, 9, and 10, Calcutta, 2000.

    Techniques Used:

    20) Product Images from "Isolation of a Novel Bacteriophage Specific for the Periodontal Pathogen Fusobacterium nucleatum ▿"

    Article Title: Isolation of a Novel Bacteriophage Specific for the Periodontal Pathogen Fusobacterium nucleatum ▿

    Journal: Applied and Environmental Microbiology

    doi: 10.1128/AEM.01135-10

    Restriction digest patterns electrophoresed on a 1.5% agarose gel and stained with ethidium bromide. (A) FnpΦ02 genomic DNA digested with restriction enzymes. Lanes: L1, 100-bp DNA ladder marker (Fermentas Inc., Canada); L2, 1-kb ladder marker (Fermentas Inc., Canada); L3, undigested DNA; L4, DNA/HindIII; L5, DNA/DraI; L6, DNA/XbaI; and L7, lambda DNA/HindIII. (B) Higher magnification of the bands of the digestion of FnpΦ02 with HindIII used for genome size determination. Selected sizes of the marker are indicated in panel A.
    Figure Legend Snippet: Restriction digest patterns electrophoresed on a 1.5% agarose gel and stained with ethidium bromide. (A) FnpΦ02 genomic DNA digested with restriction enzymes. Lanes: L1, 100-bp DNA ladder marker (Fermentas Inc., Canada); L2, 1-kb ladder marker (Fermentas Inc., Canada); L3, undigested DNA; L4, DNA/HindIII; L5, DNA/DraI; L6, DNA/XbaI; and L7, lambda DNA/HindIII. (B) Higher magnification of the bands of the digestion of FnpΦ02 with HindIII used for genome size determination. Selected sizes of the marker are indicated in panel A.

    Techniques Used: Agarose Gel Electrophoresis, Staining, Marker, Lambda DNA Preparation

    21) Product Images from "Engineering BioBrick vectors from BioBrick parts"

    Article Title: Engineering BioBrick vectors from BioBrick parts

    Journal: Journal of Biological Engineering

    doi: 10.1186/1754-1611-2-5

    Using the new BioBrick vectors . To verify the function of the new BioBrick vectors, we performed a colony PCR using primers that anneal to the verification primer binding sites. To check the length of the resulting PCR products, we electrophoresed the reactions through an 0.8% agarose gel. Lanes 1–8 are the PCR products resulting from the amplification of the following BioBrick parts cloned into new BioBrick vectors. The desired PCR product lengths are in parentheses. Lane 1 is pSB4A5-I52001 (1370 bp), lane 2 is pSB4K5-T9003 (1883 bp), lane 3 is pSB4C5-E0435 (814 bp), lane 4 is pSB4T5-P20061 (2988 bp), lane 5 is pSB3K5-I52002 (1370 bp), lane 6 is pSB3C5-I52001 (1370 bp), lane 7 is pSB3T5-I6413 (867 bp), and lane 8 is BBa_I51020 (1370 bp). Lane 9 is 1 μ g of 2-log DNA ladder (New England Biolabs, Inc.). The 0.5 kb, 1 kb, and 3 kb DNA fragments in the DNA ladder are annotated.
    Figure Legend Snippet: Using the new BioBrick vectors . To verify the function of the new BioBrick vectors, we performed a colony PCR using primers that anneal to the verification primer binding sites. To check the length of the resulting PCR products, we electrophoresed the reactions through an 0.8% agarose gel. Lanes 1–8 are the PCR products resulting from the amplification of the following BioBrick parts cloned into new BioBrick vectors. The desired PCR product lengths are in parentheses. Lane 1 is pSB4A5-I52001 (1370 bp), lane 2 is pSB4K5-T9003 (1883 bp), lane 3 is pSB4C5-E0435 (814 bp), lane 4 is pSB4T5-P20061 (2988 bp), lane 5 is pSB3K5-I52002 (1370 bp), lane 6 is pSB3C5-I52001 (1370 bp), lane 7 is pSB3T5-I6413 (867 bp), and lane 8 is BBa_I51020 (1370 bp). Lane 9 is 1 μ g of 2-log DNA ladder (New England Biolabs, Inc.). The 0.5 kb, 1 kb, and 3 kb DNA fragments in the DNA ladder are annotated.

    Techniques Used: Polymerase Chain Reaction, Binding Assay, Agarose Gel Electrophoresis, Amplification, Clone Assay

    22) Product Images from "Efficient Genome Editing in Clostridium cellulolyticum via CRISPR-Cas9 Nickase"

    Article Title: Efficient Genome Editing in Clostridium cellulolyticum via CRISPR-Cas9 Nickase

    Journal: Applied and Environmental Microbiology

    doi: 10.1128/AEM.00873-15

    Precise deletion and insertion of a small fragment. (A) Schematic all-in-one vector for pyrF disruption by a single-nick-triggered homologous recombination (SNHR). The vector consists of an Fd-driven cas9n gene, a P4-driven gRNA-targeting pyrF gene, and a donor template with a 23-bp deletion flanked by 1-kb left homologous (LH) and right homologous (RH) arms. (B) DNA sequence chromatograms showing the deletion of a 23-bp target site in the pyrF gene. The 23-bp region carries a 20-base gRNA sequence and a 3-base protospacer adjacent motif (PAM). Twelve colonies all present precise deletion at the position indicated by a downward black arrow. Amplicon for sequencing was generated using primers p1 and p2, as schematized in panel A. (C) SNHR-mediated insertion of an EcoRV site at a target cut site in the β- gal gene. The donor template shown in the dashed box carries the EcoRV site flanked by 1-kb LH and RH starting from the Cas9n cleavage site. (D) PCR identification of Δ gal mutants. The transformant population of empty vector (CK) and pCas9n-β- gal -donor (R1 and R2, two replicates) is identified by two primer pairs as drawn in panel C. (E) EcoRV digestion of p3/p4 PCR products. (F) DNA sequence chromatograms verifying the precise insertion of EcoRV (underlined) in the Δβ- gal mutant.
    Figure Legend Snippet: Precise deletion and insertion of a small fragment. (A) Schematic all-in-one vector for pyrF disruption by a single-nick-triggered homologous recombination (SNHR). The vector consists of an Fd-driven cas9n gene, a P4-driven gRNA-targeting pyrF gene, and a donor template with a 23-bp deletion flanked by 1-kb left homologous (LH) and right homologous (RH) arms. (B) DNA sequence chromatograms showing the deletion of a 23-bp target site in the pyrF gene. The 23-bp region carries a 20-base gRNA sequence and a 3-base protospacer adjacent motif (PAM). Twelve colonies all present precise deletion at the position indicated by a downward black arrow. Amplicon for sequencing was generated using primers p1 and p2, as schematized in panel A. (C) SNHR-mediated insertion of an EcoRV site at a target cut site in the β- gal gene. The donor template shown in the dashed box carries the EcoRV site flanked by 1-kb LH and RH starting from the Cas9n cleavage site. (D) PCR identification of Δ gal mutants. The transformant population of empty vector (CK) and pCas9n-β- gal -donor (R1 and R2, two replicates) is identified by two primer pairs as drawn in panel C. (E) EcoRV digestion of p3/p4 PCR products. (F) DNA sequence chromatograms verifying the precise insertion of EcoRV (underlined) in the Δβ- gal mutant.

    Techniques Used: Plasmid Preparation, Homologous Recombination, Sequencing, Amplification, Generated, Polymerase Chain Reaction, Mutagenesis

    Evaluation of editing efficacy and cargo capacity. (A and B) Effect of arm size on editing efficacy. (A) Design of donor templates with various arm sizes (0.1 to 1 kb), in which the target site (red) is modified to contain an EcoRV site (yellow). The all-in-one vectors with these templates introduce EcoRV into the β- gal gene via SNHR. (B) Editing efficacy evaluation by EcoRV digestion of p3/p4 PCR product. The percentage of edited genome in the whole population of control with donor-free vector (CK), recovered cells (T0), and TMP-resistant cells from three serial transfers (T1 to T3) is calculated by densitometry analysis. (C to F) Genetic cargo capacity evaluation by delivering foreign DNA fragments with various sizes into the genome. (C) Design of four donor templates with 0.71-kb Fd:: afp , 1.72-kb promoterless alsS , and 3-kb and 6-kb λ DNA (blue) between 1-kb arms. Using SNHR, the alsS fragment and the remainder are inserted into two different sites, 3198D and pyrF , respectively. (D) PCR identification of Δ pyrF/afp + and alsS + mutants generated by the insertion of Fd:: afp and alsS fragments, using wild type (CK) as a control. Primer pairs are indicated and drawn in panel C. (E) Enrichment of Δ pyrF/afp + mutant in the population during serial transfer (T0 to T3) using wild type (CK) as control. (F) Fluorescence microscopy of plasmid-cured Δ pyrF/afp + mutant.
    Figure Legend Snippet: Evaluation of editing efficacy and cargo capacity. (A and B) Effect of arm size on editing efficacy. (A) Design of donor templates with various arm sizes (0.1 to 1 kb), in which the target site (red) is modified to contain an EcoRV site (yellow). The all-in-one vectors with these templates introduce EcoRV into the β- gal gene via SNHR. (B) Editing efficacy evaluation by EcoRV digestion of p3/p4 PCR product. The percentage of edited genome in the whole population of control with donor-free vector (CK), recovered cells (T0), and TMP-resistant cells from three serial transfers (T1 to T3) is calculated by densitometry analysis. (C to F) Genetic cargo capacity evaluation by delivering foreign DNA fragments with various sizes into the genome. (C) Design of four donor templates with 0.71-kb Fd:: afp , 1.72-kb promoterless alsS , and 3-kb and 6-kb λ DNA (blue) between 1-kb arms. Using SNHR, the alsS fragment and the remainder are inserted into two different sites, 3198D and pyrF , respectively. (D) PCR identification of Δ pyrF/afp + and alsS + mutants generated by the insertion of Fd:: afp and alsS fragments, using wild type (CK) as a control. Primer pairs are indicated and drawn in panel C. (E) Enrichment of Δ pyrF/afp + mutant in the population during serial transfer (T0 to T3) using wild type (CK) as control. (F) Fluorescence microscopy of plasmid-cured Δ pyrF/afp + mutant.

    Techniques Used: Modification, Introduce, Polymerase Chain Reaction, Plasmid Preparation, Generated, Mutagenesis, Fluorescence, Microscopy

    Bioinformatic analysis of targeting space in  C. cellulolyticum . (A) Genome-wide distribution of genes and target sites on both DNA strands. White areas in each track indicate gaps between adjacent genes or target sites. The color code is given below the map. (B) Histogram of distance between adjacent usable target sites. Values of mean and median, the number of untouchable regions (UR) with lengths of  > 1 kb, and the length of the maximal UR are inset within the plot. (C) Histogram of the number of usable target sites in genes. The values of mean, median, and gene coverage are inset.
    Figure Legend Snippet: Bioinformatic analysis of targeting space in C. cellulolyticum . (A) Genome-wide distribution of genes and target sites on both DNA strands. White areas in each track indicate gaps between adjacent genes or target sites. The color code is given below the map. (B) Histogram of distance between adjacent usable target sites. Values of mean and median, the number of untouchable regions (UR) with lengths of > 1 kb, and the length of the maximal UR are inset within the plot. (C) Histogram of the number of usable target sites in genes. The values of mean, median, and gene coverage are inset.

    Techniques Used: Genome Wide

    23) Product Images from "Identification of RNase-resistant RNAs in Saccharomyces cerevisiae extracts: separation from chromosomal DNA by selective precipitation"

    Article Title: Identification of RNase-resistant RNAs in Saccharomyces cerevisiae extracts: separation from chromosomal DNA by selective precipitation

    Journal: Analytical biochemistry

    doi: 10.1016/j.ab.2015.09.017

    The lower and upper RNase-resistant bands are composed of RNA fragments ranging in size from 10-16 bp and 24-56 bp, respectively. (A) Lane 1, S. cerevisiae chromosomal DNA treated with RNase A was run on a 3.5% agarose gel; M, dsRNA ladder; (B) The logs
    Figure Legend Snippet: The lower and upper RNase-resistant bands are composed of RNA fragments ranging in size from 10-16 bp and 24-56 bp, respectively. (A) Lane 1, S. cerevisiae chromosomal DNA treated with RNase A was run on a 3.5% agarose gel; M, dsRNA ladder; (B) The logs

    Techniques Used: Agarose Gel Electrophoresis

    Treatment of chromosomal DNA minipreps with potassium acetate and isopropanol removes the RNase-resistant RNA fragments. All samples were run on 1.2% agarose gels. (A) Representative examples of treatment with varying volumes of 88% isopropanol/0.2 M
    Figure Legend Snippet: Treatment of chromosomal DNA minipreps with potassium acetate and isopropanol removes the RNase-resistant RNA fragments. All samples were run on 1.2% agarose gels. (A) Representative examples of treatment with varying volumes of 88% isopropanol/0.2 M

    Techniques Used:

    Small RNase A-resistant bands are retained in yeast chromosomal DNA minipreps but not in E. coli bacterial chromosomal DNA minipreps. All DNA samples were run on 1.2% agarose gels. (A) Lanes 1 and 2, S. cerevisiae chromosomal DNA extracted using a simple
    Figure Legend Snippet: Small RNase A-resistant bands are retained in yeast chromosomal DNA minipreps but not in E. coli bacterial chromosomal DNA minipreps. All DNA samples were run on 1.2% agarose gels. (A) Lanes 1 and 2, S. cerevisiae chromosomal DNA extracted using a simple

    Techniques Used:

    Conventional physical and enzymatic methods were unsuccessful at removing both RNase-resistant populations from S. cerevisiae DNA minipreps. Samples were run on 1.2% agarose gels. Two bands persisted after treatment with RNase A (Lane 1 in panels A, B,
    Figure Legend Snippet: Conventional physical and enzymatic methods were unsuccessful at removing both RNase-resistant populations from S. cerevisiae DNA minipreps. Samples were run on 1.2% agarose gels. Two bands persisted after treatment with RNase A (Lane 1 in panels A, B,

    Techniques Used:

    Base hydrolysis methods eliminated the resistant RNA bands but strongly reduced the yield of chromosomal DNA. Lane 1, DNA after RNase A treatment; lane 2, DNA was digested with RNase A, followed by standard NaOAc/ethanol precipitation; lanes 3 and 4,
    Figure Legend Snippet: Base hydrolysis methods eliminated the resistant RNA bands but strongly reduced the yield of chromosomal DNA. Lane 1, DNA after RNase A treatment; lane 2, DNA was digested with RNase A, followed by standard NaOAc/ethanol precipitation; lanes 3 and 4,

    Techniques Used: Ethanol Precipitation

    24) Product Images from "Sizing femtogram amounts of dsDNA by single-molecule counting"

    Article Title: Sizing femtogram amounts of dsDNA by single-molecule counting

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkv904

    Single-molecule intensity distribution of a 1 kbp DNA ladder. ( A ) intensity distribution histogram of detected molecules (red), fitted Gaussians sum (black) and the residue of the fitting (blue). ( B ) a plot of expected DNA lengths in the ladder sample versus the centers of the fitted Gaussian peaks exhibits linear dependency with R 2 = 0.99932.
    Figure Legend Snippet: Single-molecule intensity distribution of a 1 kbp DNA ladder. ( A ) intensity distribution histogram of detected molecules (red), fitted Gaussians sum (black) and the residue of the fitting (blue). ( B ) a plot of expected DNA lengths in the ladder sample versus the centers of the fitted Gaussian peaks exhibits linear dependency with R 2 = 0.99932.

    Techniques Used:

    25) Product Images from "The Myb/SANT domain of the telomere-binding protein TRF2 alters chromatin structure"

    Article Title: The Myb/SANT domain of the telomere-binding protein TRF2 alters chromatin structure

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkp515

    Analysis of nucleosomal array fibers. The pRST5 plasmid and expected fragments created by PvuII or SfaNI digestion ( A ). Micrococcal nuclease digestion at indicated time points of nucleosomal arrays reconstituted onto PvuII ( B ); SfaNI ( C ) digested pRST5 DNA. Multi-gels of telomeric nucleosomal array fibers (NA) and histone-free DNA (DNA) from pRST5 digested with PvuII ( D ); and SfaNI ( E ) prepared and subjected to electrophoresis according to ‘Materials and Methods’ section. Spheres refer to carboxylate-coated microsphere standards (35 nm radius). The ‘1 kb tel’ and ‘2 kb tel’ refer to the telomeric fragments liberated by PvuII and SfaNI digestion respectively. ‘N’ refers to the fragments without telomeric DNA. Logarithmic plot of pore sizes ( P e ) versus agarose% ( F ). Data was obtained from multi-gels run with bacteriophage T3 (Phage) in this laboratory or previous work [Phage, previous, (  24 )] and 35 nm carboxylate-coated microspheres (Microspheres).  P e  for each agarose concentration was calculated according to ‘Materials and Methods’. Symbols with error bars represent the mean ± 1 SD of four to eight determinations.
    Figure Legend Snippet: Analysis of nucleosomal array fibers. The pRST5 plasmid and expected fragments created by PvuII or SfaNI digestion ( A ). Micrococcal nuclease digestion at indicated time points of nucleosomal arrays reconstituted onto PvuII ( B ); SfaNI ( C ) digested pRST5 DNA. Multi-gels of telomeric nucleosomal array fibers (NA) and histone-free DNA (DNA) from pRST5 digested with PvuII ( D ); and SfaNI ( E ) prepared and subjected to electrophoresis according to ‘Materials and Methods’ section. Spheres refer to carboxylate-coated microsphere standards (35 nm radius). The ‘1 kb tel’ and ‘2 kb tel’ refer to the telomeric fragments liberated by PvuII and SfaNI digestion respectively. ‘N’ refers to the fragments without telomeric DNA. Logarithmic plot of pore sizes ( P e ) versus agarose% ( F ). Data was obtained from multi-gels run with bacteriophage T3 (Phage) in this laboratory or previous work [Phage, previous, ( 24 )] and 35 nm carboxylate-coated microspheres (Microspheres). P e for each agarose concentration was calculated according to ‘Materials and Methods’. Symbols with error bars represent the mean ± 1 SD of four to eight determinations.

    Techniques Used: Plasmid Preparation, Electrophoresis, Concentration Assay

    TRF2 DBD binds specifically to telomeric DNA and nucleosomal arrays. 0.6% agarose gels of TRF2 DBD binding to DNA (DNA) ( A ) and nucleosomal arrays (NA) ( B ) from the pRST5 fragment digested to obtain a 1-kb DNA fragment with 580-bp telomeric DNA (telo) and 2.5-kb non-telomeric DNA (N-telo). Gels similar to (A) and (B), respectively except the pRST5 was digested to obtain a 2-kb fragment containing the 580-bp telomeric DNA (telo) with a 1 kb and smaller fragments being non-telomeric (N-telo) ( C and D ).
    Figure Legend Snippet: TRF2 DBD binds specifically to telomeric DNA and nucleosomal arrays. 0.6% agarose gels of TRF2 DBD binding to DNA (DNA) ( A ) and nucleosomal arrays (NA) ( B ) from the pRST5 fragment digested to obtain a 1-kb DNA fragment with 580-bp telomeric DNA (telo) and 2.5-kb non-telomeric DNA (N-telo). Gels similar to (A) and (B), respectively except the pRST5 was digested to obtain a 2-kb fragment containing the 580-bp telomeric DNA (telo) with a 1 kb and smaller fragments being non-telomeric (N-telo) ( C and D ).

    Techniques Used: Binding Assay

    26) Product Images from "An Optimized Protocol for ChIP-Seq from Human Embryonic Stem Cell Cultures"

    Article Title: An Optimized Protocol for ChIP-Seq from Human Embryonic Stem Cell Cultures

    Journal: Star Protocols

    doi: 10.1016/j.xpro.2020.100062

    An Example of a Sonication Optimization Experiment, Where Fixed H1 hESCs Were Sheared Using a Bioruptor Plus Sonication System for the Indicated Number of Cycles of 30 s ON/30 s OFF Sheared DNA was then purified and separated on a 1.8% Agarose Gel. 10 was determined to be the minimum number of cycles required to shear the majority of chromatin into the 100 bp to 500 bp range.
    Figure Legend Snippet: An Example of a Sonication Optimization Experiment, Where Fixed H1 hESCs Were Sheared Using a Bioruptor Plus Sonication System for the Indicated Number of Cycles of 30 s ON/30 s OFF Sheared DNA was then purified and separated on a 1.8% Agarose Gel. 10 was determined to be the minimum number of cycles required to shear the majority of chromatin into the 100 bp to 500 bp range.

    Techniques Used: Sonication, Purification, Agarose Gel Electrophoresis

    Example Sonicated Chromatin (Input Samples) from H1 hESC Cultured in mTeSR1 on Matrigel, Separated on an Agilent Tapestation System with a High Sensitivity D1000 ScreenTape Both the pseudo-gel image (A) and the plot of signal intensity vs predicted fragment size (B) show that the majority of the DNA lies in the 100 bp to 500 bp range, with an intensity peak at 260 bp and an average size of 352 bp for signal over the 50–1,000 bp range. Upper (1,500 bp) and Lower (25 bp) size markers are indicated.
    Figure Legend Snippet: Example Sonicated Chromatin (Input Samples) from H1 hESC Cultured in mTeSR1 on Matrigel, Separated on an Agilent Tapestation System with a High Sensitivity D1000 ScreenTape Both the pseudo-gel image (A) and the plot of signal intensity vs predicted fragment size (B) show that the majority of the DNA lies in the 100 bp to 500 bp range, with an intensity peak at 260 bp and an average size of 352 bp for signal over the 50–1,000 bp range. Upper (1,500 bp) and Lower (25 bp) size markers are indicated.

    Techniques Used: Sonication, Cell Culture

    27) Product Images from "Instability of the Octarepeat Region of the Human Prion Protein Gene"

    Article Title: Instability of the Octarepeat Region of the Human Prion Protein Gene

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0026635

    Mutant plasmids from octarepeat replication in DH5α are all head-to-head dimers. (A) Restriction analysis of replication-mutant plasmids with Spe I, Sac II and Sca I. pOct5 or pOct11b were transformed into DH5α. Plasmid DNAs were prepared from two mutant colonies and two control (non-mutant) colonies each for pOct5 and pOct11b, digested with Spe I, Sac II or Sca I, and separated by agarose gel electrophoresis. All mutant colonies appear to contain a minute amount of monomer plasmids. M1, 100-bp DNA ladder; M2, 1-kb DNA ladder. (B) Diagram of the head-to-head plasmid dimers. The top panel depicts the parental plasmid monomer; the bottom panel depicts the dimer where the newly generated monomer unit is highlighted in thicker lines. The boxes denote the octarepeat inserts.
    Figure Legend Snippet: Mutant plasmids from octarepeat replication in DH5α are all head-to-head dimers. (A) Restriction analysis of replication-mutant plasmids with Spe I, Sac II and Sca I. pOct5 or pOct11b were transformed into DH5α. Plasmid DNAs were prepared from two mutant colonies and two control (non-mutant) colonies each for pOct5 and pOct11b, digested with Spe I, Sac II or Sca I, and separated by agarose gel electrophoresis. All mutant colonies appear to contain a minute amount of monomer plasmids. M1, 100-bp DNA ladder; M2, 1-kb DNA ladder. (B) Diagram of the head-to-head plasmid dimers. The top panel depicts the parental plasmid monomer; the bottom panel depicts the dimer where the newly generated monomer unit is highlighted in thicker lines. The boxes denote the octarepeat inserts.

    Techniques Used: Mutagenesis, Transformation Assay, Plasmid Preparation, Agarose Gel Electrophoresis, Generated

    28) Product Images from "Human PSF concentrates DNA and stimulates duplex capture in DMC1-mediated homologous pairing"

    Article Title: Human PSF concentrates DNA and stimulates duplex capture in DMC1-mediated homologous pairing

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkr1229

    The DNA aggregation assay. ( A ) Schematic representation of the DNA aggregation assay. ( B ) The reaction was conducted with DMC1 (4 µM) and/or PSF (1.2 µM) in the presence of ϕX174 ssDNA (10 µM) and linearized ϕX174 dsDNA (10 µM). The samples were centrifuged for 3 min at 20 400 g at room temperature, and the ssDNA and dsDNA recovered in the upper (15 µl) and bottom (5 µl) fractions were analyzed by 0.8% agarose gel electrophoresis with ethidium bromide staining. ( C ) The reaction was conducted by the same method as in panel B, except HOP2-MND1 (1.2 µM) was used instead of PSF.
    Figure Legend Snippet: The DNA aggregation assay. ( A ) Schematic representation of the DNA aggregation assay. ( B ) The reaction was conducted with DMC1 (4 µM) and/or PSF (1.2 µM) in the presence of ϕX174 ssDNA (10 µM) and linearized ϕX174 dsDNA (10 µM). The samples were centrifuged for 3 min at 20 400 g at room temperature, and the ssDNA and dsDNA recovered in the upper (15 µl) and bottom (5 µl) fractions were analyzed by 0.8% agarose gel electrophoresis with ethidium bromide staining. ( C ) The reaction was conducted by the same method as in panel B, except HOP2-MND1 (1.2 µM) was used instead of PSF.

    Techniques Used: Agarose Gel Electrophoresis, Staining

    29) Product Images from "In-Fusion BioBrick assembly and re-engineering"

    Article Title: In-Fusion BioBrick assembly and re-engineering

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkq179

    Part swapping: simultaneous promoter and RBS re-engineering. ( a ) R0011+E0240 and J04450 plasmids are shown with the forward and reverse primers for PCR. R0011+E0240 is amplified with the vector and J04450 is amplified as the insert. ( b ) Detailed schematic of the assembly strategy with the forward and reverse primers. Only the promoter (R0011) and RBS (B0032) are PCR-amplified from the R0011+E0240 plasmid. Only E1010 and B0010/12 are PCR-amplified from the J04450 circuit in order to change its promoter and RBS in one assembly step. ( c ) Since both plasmids used as template DNA in the PCR reaction were approximately the same size as the desired construct, two colony PCR reactions were performed on the same six colonies. The gel on the left shows six colonies amplified with VF2/VR primers and the gel on the right shows the same six colonies (#1–6) and negative control J04450 plasmid (#7) amplified with the VF2 and R0011+E0240 AR primer. Correct colonies show a PCR product of about 1.1 kb for the left gel and a PCR product of about 300 bp for the right gel (correct size indicated by arrow).
    Figure Legend Snippet: Part swapping: simultaneous promoter and RBS re-engineering. ( a ) R0011+E0240 and J04450 plasmids are shown with the forward and reverse primers for PCR. R0011+E0240 is amplified with the vector and J04450 is amplified as the insert. ( b ) Detailed schematic of the assembly strategy with the forward and reverse primers. Only the promoter (R0011) and RBS (B0032) are PCR-amplified from the R0011+E0240 plasmid. Only E1010 and B0010/12 are PCR-amplified from the J04450 circuit in order to change its promoter and RBS in one assembly step. ( c ) Since both plasmids used as template DNA in the PCR reaction were approximately the same size as the desired construct, two colony PCR reactions were performed on the same six colonies. The gel on the left shows six colonies amplified with VF2/VR primers and the gel on the right shows the same six colonies (#1–6) and negative control J04450 plasmid (#7) amplified with the VF2 and R0011+E0240 AR primer. Correct colonies show a PCR product of about 1.1 kb for the left gel and a PCR product of about 300 bp for the right gel (correct size indicated by arrow).

    Techniques Used: Polymerase Chain Reaction, Amplification, Plasmid Preparation, Construct, Negative Control

    30) Product Images from "Novel ?-Lactamase Genes from Two Environmental Isolates of Vibrio harveyi"

    Article Title: Novel ?-Lactamase Genes from Two Environmental Isolates of Vibrio harveyi

    Journal: Antimicrobial Agents and Chemotherapy

    doi:

    (A) PFGE of Not I-digested DNA from nine environmental isolates of V. harveyi . (B) Southern hybridization analysis of DNA. The probe used was the 1.1-kb Hin dIII fragment containing bla VHW-1 . Lanes: 1, W3B; 2, GCB; 3, AP5; 4, HB3; 5, R. sphaeroides 2.4.1 DNA digested with Ase I (molecular size standard); 6, AP6; 7, P1B; 8, M 1 ; 9, E 2 ; 10, M3.4L.
    Figure Legend Snippet: (A) PFGE of Not I-digested DNA from nine environmental isolates of V. harveyi . (B) Southern hybridization analysis of DNA. The probe used was the 1.1-kb Hin dIII fragment containing bla VHW-1 . Lanes: 1, W3B; 2, GCB; 3, AP5; 4, HB3; 5, R. sphaeroides 2.4.1 DNA digested with Ase I (molecular size standard); 6, AP6; 7, P1B; 8, M 1 ; 9, E 2 ; 10, M3.4L.

    Techniques Used: Hybridization

    31) Product Images from "A single catalytic domain of the junction-resolving enzyme T7 endonuclease I is a non-specific nicking endonuclease"

    Article Title: A single catalytic domain of the junction-resolving enzyme T7 endonuclease I is a non-specific nicking endonuclease

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gki921

    ( A ) Determination of non-specific nuclease activity of SCD protein. Variable amounts of purified MEn–In/Ic–Ec were incubated with 1 µg of 2-log DNA ladder (NEB) in 20 µl of either Mg 2+ or Mn 2+ buffer at 37°C for 1 h. The digests were resolved on a 1.2% agarose gel. From lane 1 to 5, digests with 0.00, 0.125, 0.25, 0.5 and 1.0 µg of MEn–In/Ic–Ec in Mg 2+ buffer, respectively. From lane 6 to 10, digests are the same as from lane 1 to 5 except for using Mn 2+ buffer. ( B ) Identification of nicked strands as intermediate products of reaction by SCD protein. Plasmid pUC19 was incubated with MEn–In/Ic–Ec in Mg 2+ buffer at 37°C. At variable time point 0, 2, 5, 10, 20, 30, 40, 60, 120 and 180 min an aliquot of sample was withdrawn from the reaction and resolved on an agarose gel. L, N and S stand for linear, nicked and supercoiled plasmids, respectively.
    Figure Legend Snippet: ( A ) Determination of non-specific nuclease activity of SCD protein. Variable amounts of purified MEn–In/Ic–Ec were incubated with 1 µg of 2-log DNA ladder (NEB) in 20 µl of either Mg 2+ or Mn 2+ buffer at 37°C for 1 h. The digests were resolved on a 1.2% agarose gel. From lane 1 to 5, digests with 0.00, 0.125, 0.25, 0.5 and 1.0 µg of MEn–In/Ic–Ec in Mg 2+ buffer, respectively. From lane 6 to 10, digests are the same as from lane 1 to 5 except for using Mn 2+ buffer. ( B ) Identification of nicked strands as intermediate products of reaction by SCD protein. Plasmid pUC19 was incubated with MEn–In/Ic–Ec in Mg 2+ buffer at 37°C. At variable time point 0, 2, 5, 10, 20, 30, 40, 60, 120 and 180 min an aliquot of sample was withdrawn from the reaction and resolved on an agarose gel. L, N and S stand for linear, nicked and supercoiled plasmids, respectively.

    Techniques Used: Activity Assay, Purification, Incubation, Agarose Gel Electrophoresis, Plasmid Preparation

    Determination of the ratio of structure-specific to non-specific activity of SCD protein by agarose gel-electrophoresis. ( A ) Schematic illustration of possible linearization sites on pUC(AT) by both specific and non-specific activities of SCD protein. The long arrow represents the specific activity (Sp) that leads the plasmid to open at the cruciform site. The short arrows represent the non-specific activity (Non sp) that leads the plasmid to open at variable sites. ( B ) LP and SP stand for linear and supercoiled form plasmids, respectively; LF and SF for the large and the small fragments produced by DrdI digestion respectively. Lane 1, pUC(AT); lane 2, pUC(AT) cut by DrdI (a small amount of linear plasmid was produced by incomplete digestion); lane 3, linear pUC(AT) produced by T7 Endo I; lane 4, the DNA in lane 3 cut by DrdI; lane 5, linear pUC(AT) produced by MEn–In/Ic–Ec; lane 6, the DNA in lane 5 cut by DrdI; lane 7, linear pUC(AT) produced by MEn–In(9)/Ic–Ec; lane 8, the DNA in lane 7 cut by DrdI; lane 9, linear pUC19 produced by MEn–In/Ic–Ec (a small amount of supercoiled plasmid is co-purified with the linear form); lane 10, the DNA in lane 9 cut by DrdI. Each lane contained ∼1 µg of DNA. All the linear plasmids used in the assay were gel-purified.
    Figure Legend Snippet: Determination of the ratio of structure-specific to non-specific activity of SCD protein by agarose gel-electrophoresis. ( A ) Schematic illustration of possible linearization sites on pUC(AT) by both specific and non-specific activities of SCD protein. The long arrow represents the specific activity (Sp) that leads the plasmid to open at the cruciform site. The short arrows represent the non-specific activity (Non sp) that leads the plasmid to open at variable sites. ( B ) LP and SP stand for linear and supercoiled form plasmids, respectively; LF and SF for the large and the small fragments produced by DrdI digestion respectively. Lane 1, pUC(AT); lane 2, pUC(AT) cut by DrdI (a small amount of linear plasmid was produced by incomplete digestion); lane 3, linear pUC(AT) produced by T7 Endo I; lane 4, the DNA in lane 3 cut by DrdI; lane 5, linear pUC(AT) produced by MEn–In/Ic–Ec; lane 6, the DNA in lane 5 cut by DrdI; lane 7, linear pUC(AT) produced by MEn–In(9)/Ic–Ec; lane 8, the DNA in lane 7 cut by DrdI; lane 9, linear pUC19 produced by MEn–In/Ic–Ec (a small amount of supercoiled plasmid is co-purified with the linear form); lane 10, the DNA in lane 9 cut by DrdI. Each lane contained ∼1 µg of DNA. All the linear plasmids used in the assay were gel-purified.

    Techniques Used: Activity Assay, Agarose Gel Electrophoresis, Plasmid Preparation, Produced, Purification

    32) Product Images from "BRD4 inhibitors block telomere elongation"

    Article Title: BRD4 inhibitors block telomere elongation

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkx561

    Three additional BRD4 inhibitors block telomere inhibition in a dose-dependent manner. Mouse fibroblasts transduced with SVA lentivirus were treated with four different BRD4 inhibitors: JQ1, IBET151, MS436 OTX015. ( A ) Southern blot of mouse fibroblast telomeric DNA, at days 2 and 6 post SVA transduction, grown in the presence of DMSO (lane 2–3), 10 μM KU-55933 (lane 4–5), 0.1 μM JQ1 (lane 6–7), 5 μM MS436 (lane 8–9), 0.5 μM OTX015 (lane 10–11) or 1 μM IBET151 (lane 12–13). ( B ) Dose dependence of IBET151. Southern blot of mouse fibroblast telomeric DNA, at days 2 and 6 post SVA transduction, in the presence of 1 μM (lane 2–3), 0.5 μM (lane 4–5), 0.25 (lane 6–7) or 0.125 μM (lane 8–9) IBET151. ( C ) Dose dependence of OTX015 and MS436. Southern blot of mouse fibroblast telomeric DNA, at days 2 and 6 post SVA transduction, in the presence of DMSO (lane 2–3), 10 μM KU-55933 (lane 4–5), 250 nM (lane 6–7), 125 nM (lane 8–9) or 62.5 nM OTX015 (lane 10–11), 5 μM (lane 12–13), 2.5 μM (lane 14–15), 1.25 μM (lane 16–17) or 0.625 μM (lane 18–19) MS436. Lane 1 in all panels shows the 2-log ladder marker (NEB), sizes marked are in kilobases. In Panel B, other lanes between the marker and IBET treated lanes were removed.
    Figure Legend Snippet: Three additional BRD4 inhibitors block telomere inhibition in a dose-dependent manner. Mouse fibroblasts transduced with SVA lentivirus were treated with four different BRD4 inhibitors: JQ1, IBET151, MS436 OTX015. ( A ) Southern blot of mouse fibroblast telomeric DNA, at days 2 and 6 post SVA transduction, grown in the presence of DMSO (lane 2–3), 10 μM KU-55933 (lane 4–5), 0.1 μM JQ1 (lane 6–7), 5 μM MS436 (lane 8–9), 0.5 μM OTX015 (lane 10–11) or 1 μM IBET151 (lane 12–13). ( B ) Dose dependence of IBET151. Southern blot of mouse fibroblast telomeric DNA, at days 2 and 6 post SVA transduction, in the presence of 1 μM (lane 2–3), 0.5 μM (lane 4–5), 0.25 (lane 6–7) or 0.125 μM (lane 8–9) IBET151. ( C ) Dose dependence of OTX015 and MS436. Southern blot of mouse fibroblast telomeric DNA, at days 2 and 6 post SVA transduction, in the presence of DMSO (lane 2–3), 10 μM KU-55933 (lane 4–5), 250 nM (lane 6–7), 125 nM (lane 8–9) or 62.5 nM OTX015 (lane 10–11), 5 μM (lane 12–13), 2.5 μM (lane 14–15), 1.25 μM (lane 16–17) or 0.625 μM (lane 18–19) MS436. Lane 1 in all panels shows the 2-log ladder marker (NEB), sizes marked are in kilobases. In Panel B, other lanes between the marker and IBET treated lanes were removed.

    Techniques Used: Blocking Assay, Inhibition, Transduction, Southern Blot, Marker

    JQ1 blocks telomere elongation in a dose-dependent manner. Mouse fibroblasts were transduced with SVA lentivirus, encoding for mTERT and mTR and cultured for six days. ( A ) Southern blot of mouse fibroblast telomeric DNA, at days 2 and 6 post-SVA transduction, grown in the presence of DMSO (lane 2–3), 10 μM KU-55933 (lane 4–5) or 0.1 μM JQ1 (lane 6–7). Lane 1 shows the 2-log ladder marker (NEB) sizes marked in kilobases. ( B ) Southern blot of mouse fibroblast telomeric DNA, at days 2 and 6 post-SVA transduction, in the presence of DMSO (lane 2–3), 10 μM KU-55933 (lane 4–5) or decreasing concentrations of 33 nM JQ1 (lane 6–7), 11 nM JQ1 (lane 8–9) or 3.3 nM JQ1 (lane 10–11). Lane 1 shows the 2-log ladder marker (NEB) sizes marked are in kilobases.
    Figure Legend Snippet: JQ1 blocks telomere elongation in a dose-dependent manner. Mouse fibroblasts were transduced with SVA lentivirus, encoding for mTERT and mTR and cultured for six days. ( A ) Southern blot of mouse fibroblast telomeric DNA, at days 2 and 6 post-SVA transduction, grown in the presence of DMSO (lane 2–3), 10 μM KU-55933 (lane 4–5) or 0.1 μM JQ1 (lane 6–7). Lane 1 shows the 2-log ladder marker (NEB) sizes marked in kilobases. ( B ) Southern blot of mouse fibroblast telomeric DNA, at days 2 and 6 post-SVA transduction, in the presence of DMSO (lane 2–3), 10 μM KU-55933 (lane 4–5) or decreasing concentrations of 33 nM JQ1 (lane 6–7), 11 nM JQ1 (lane 8–9) or 3.3 nM JQ1 (lane 10–11). Lane 1 shows the 2-log ladder marker (NEB) sizes marked are in kilobases.

    Techniques Used: Transduction, Cell Culture, Southern Blot, Marker

    BRD4 inhibition causes telomere shortening in human and mouse cells in culture. ( A ) Southern blot of telomeric DNA from HeLa cells treated with DMSO (lane 2–5), or 2.5 μM OTX015 (lane 6–9) for 6 weeks, with samples taken at 2, 4 and 6 weeks of treatment. ( B ) Southern blot of telomeric DNA from mouse fibroblast cells, which were treated with DMSO (lane 2–5), or 0.5 μM OTX015 (lane 6–9) for 6 weeks, with samples taken at 2, 4 and 6 weeks of treatment.
    Figure Legend Snippet: BRD4 inhibition causes telomere shortening in human and mouse cells in culture. ( A ) Southern blot of telomeric DNA from HeLa cells treated with DMSO (lane 2–5), or 2.5 μM OTX015 (lane 6–9) for 6 weeks, with samples taken at 2, 4 and 6 weeks of treatment. ( B ) Southern blot of telomeric DNA from mouse fibroblast cells, which were treated with DMSO (lane 2–5), or 0.5 μM OTX015 (lane 6–9) for 6 weeks, with samples taken at 2, 4 and 6 weeks of treatment.

    Techniques Used: Inhibition, Southern Blot

    33) Product Images from "Intravesicle Isothermal DNA Replication"

    Article Title: Intravesicle Isothermal DNA Replication

    Journal: BMC Research Notes

    doi: 10.1186/1756-0500-4-128

    The effect of temperature on tHDA enzymatic activity . (A) The influence of temperature on the enzymatic activity of the isothermal tHDA system. Samples were incubated at 65°C (lane 2), 37°C (lane 3), 23°C (lane 4), and 4°C (lane 5) for 1.5 h. Lane 1 contains a 50 bp DNA ladder. The 1350 bp, 100 bp, and 50 bp bands are labeled. (B) The influence of overnight incubation temperature on the enzymatic activity of the isothermal tHDA system. Samples were incubated at 4°C (lane 2) and 23°C (lane 3) overnight prior to incubation at 65°C for 1.5 h. Lane 1 is a 50 bp DNA ladder. Reaction products were observed by ethidium bromide staining of a 1.8% agarose gel.
    Figure Legend Snippet: The effect of temperature on tHDA enzymatic activity . (A) The influence of temperature on the enzymatic activity of the isothermal tHDA system. Samples were incubated at 65°C (lane 2), 37°C (lane 3), 23°C (lane 4), and 4°C (lane 5) for 1.5 h. Lane 1 contains a 50 bp DNA ladder. The 1350 bp, 100 bp, and 50 bp bands are labeled. (B) The influence of overnight incubation temperature on the enzymatic activity of the isothermal tHDA system. Samples were incubated at 4°C (lane 2) and 23°C (lane 3) overnight prior to incubation at 65°C for 1.5 h. Lane 1 is a 50 bp DNA ladder. Reaction products were observed by ethidium bromide staining of a 1.8% agarose gel.

    Techniques Used: Activity Assay, Incubation, Labeling, Staining, Agarose Gel Electrophoresis

    The influence of freeze/thaw cycles on tHDA enzymatic activity . Unencapsulated reaction mixtures were subjected to either 0 (lane 2), 5 (lane 3), 10 (lane 4), or 20 (lane 5) cycles of freeze-thawing. Lane 1 is a 50 bp DNA ladder. The 1350 bp, 100 bp, and 50 bp bands are labeled. Reaction products were visualized by ethidium bromide staining of a 1.8% agarose gel. The full length reaction product is 85 bp.
    Figure Legend Snippet: The influence of freeze/thaw cycles on tHDA enzymatic activity . Unencapsulated reaction mixtures were subjected to either 0 (lane 2), 5 (lane 3), 10 (lane 4), or 20 (lane 5) cycles of freeze-thawing. Lane 1 is a 50 bp DNA ladder. The 1350 bp, 100 bp, and 50 bp bands are labeled. Reaction products were visualized by ethidium bromide staining of a 1.8% agarose gel. The full length reaction product is 85 bp.

    Techniques Used: Activity Assay, Labeling, Staining, Agarose Gel Electrophoresis

    Intravesicular isothermal replication . Lanes 1 and 7 contain a 50 bp DNA ladder. Lanes 2 and 3 are from unencapsulated reactions and lanes 4-6 are within phospholipid vesicles. Lanes 2 and 3 are isothermal replication reactions under standard conditions except that 0.5 mM CaCl 2 was added. CaCl 2 is needed for proteinase K activity. Further, lane 3 contained 0.9 units of proteinase K, demonstrating that proteinase K is capable of digesting components of the tHDA system and thus inhibiting DNA amplification. A similar experiment was conducted inside of vesicles with proteinase K added to the outside of the vesicles (lane 4) and proteinase K added to both the inside and the outside of the vesicles (lane 5). Finally, to further demonstrate that the reaction was encapsulated, dNTPs were added extravesicularly resulting in an inability to replicate DNA since dNTPs are incapable of crossing POPC membranes (lane 6).
    Figure Legend Snippet: Intravesicular isothermal replication . Lanes 1 and 7 contain a 50 bp DNA ladder. Lanes 2 and 3 are from unencapsulated reactions and lanes 4-6 are within phospholipid vesicles. Lanes 2 and 3 are isothermal replication reactions under standard conditions except that 0.5 mM CaCl 2 was added. CaCl 2 is needed for proteinase K activity. Further, lane 3 contained 0.9 units of proteinase K, demonstrating that proteinase K is capable of digesting components of the tHDA system and thus inhibiting DNA amplification. A similar experiment was conducted inside of vesicles with proteinase K added to the outside of the vesicles (lane 4) and proteinase K added to both the inside and the outside of the vesicles (lane 5). Finally, to further demonstrate that the reaction was encapsulated, dNTPs were added extravesicularly resulting in an inability to replicate DNA since dNTPs are incapable of crossing POPC membranes (lane 6).

    Techniques Used: Activity Assay, Amplification

    34) Product Images from "Defining synonymous codon compression schemes by genome recoding"

    Article Title: Defining synonymous codon compression schemes by genome recoding

    Journal: Nature

    doi: 10.1038/nature20124

    REXER enables site-specific integration of large DNA fragments into the genome. a . The use of two distinct double selection cassettes -1/+1 ( rpsL-Kan R ) and -2/+2 ( sacB-Cm R ) allows for simultaneous selection for the loss of the negative selection marker on the genome and the gain of the positive selection marker from the BAC, upon integration of synthetic DNA. b . Efficient replacement of genomic rpsL-Kan R with BAC bound sacB-Cm R using REXER 2 and REXER 4. All colonies contained the correct combination of selection markers after REXER 2 or REXER 4 as analysed by phenotyping, colony PCR, and DNA sequencing (not shown) (n = 22). c . Efficient insertion of 9 kb synthetic DNA. Genomic rpsL-Kan R was replaced with a synthetic lux operon coupled to sacB-Cm R using REXER 2 and REXER 4. All colonies on the 10-fold dilution double selection plates for REXER 2 and the 10 4 -fold plates for REXER 4 show bioluminescence. 11 colonies each from REXER 2 and REXER 4 showed correct integration by phenotyping, colony PCR, and DNA sequencing (not shown). d. Efficient insertion of 90kb synthetic DNA. The 90 kb DNA consisted of the lux operon in the middle of 80 kb DNA (previously deleted from the MDS42 genome) and followed by sacB-Cm R .
    Figure Legend Snippet: REXER enables site-specific integration of large DNA fragments into the genome. a . The use of two distinct double selection cassettes -1/+1 ( rpsL-Kan R ) and -2/+2 ( sacB-Cm R ) allows for simultaneous selection for the loss of the negative selection marker on the genome and the gain of the positive selection marker from the BAC, upon integration of synthetic DNA. b . Efficient replacement of genomic rpsL-Kan R with BAC bound sacB-Cm R using REXER 2 and REXER 4. All colonies contained the correct combination of selection markers after REXER 2 or REXER 4 as analysed by phenotyping, colony PCR, and DNA sequencing (not shown) (n = 22). c . Efficient insertion of 9 kb synthetic DNA. Genomic rpsL-Kan R was replaced with a synthetic lux operon coupled to sacB-Cm R using REXER 2 and REXER 4. All colonies on the 10-fold dilution double selection plates for REXER 2 and the 10 4 -fold plates for REXER 4 show bioluminescence. 11 colonies each from REXER 2 and REXER 4 showed correct integration by phenotyping, colony PCR, and DNA sequencing (not shown). d. Efficient insertion of 90kb synthetic DNA. The 90 kb DNA consisted of the lux operon in the middle of 80 kb DNA (previously deleted from the MDS42 genome) and followed by sacB-Cm R .

    Techniques Used: Selection, Marker, BAC Assay, Polymerase Chain Reaction, DNA Sequencing

    Simultaneous double selection and recombination enhances integration at a target locus. a. Classical recombination and double selection recombination. In classical recombination, a linear double stranded DNA with a synthetic DNA (s. DNA) sequence and a positive selection marker (+, Cm R ) flanked by homologous region 1 (HR1) and homologous region 2 (HR2) is transformed into the cell. Recombinants are selected by expression of the positive selection marker. By simultaneous double selection recombination, s. DNA containing double selection marker -2/+2 ( sacB-Cm R ) is integrated in place of the double selection marker -1/+1 ( rpsL - Kan R ) on the genome. Double selection for the gain of +2 and loss of -1 selects for simultaneous gain of s. DNA and loss of genomic sequence, and improves recombination at the target genomic locus. b. Colony PCR of clones from classical recombination and simultaneous double selection and recombination. c. All of the clones isolated by simultaneous double selection and recombination have s. DNA integrated at the target locus. The data show the mean of three independent experiments, the error bars represent the standard deviation (n=6). d . Both simultaneous double selection recombination (n = 8), and REXER 2 and REXER 4 (n = 296) result in the right combination of markers. A previously reported method integrating foreign DNA into B. subtilis , . A previously reported method replacing S. cerevisiae .
    Figure Legend Snippet: Simultaneous double selection and recombination enhances integration at a target locus. a. Classical recombination and double selection recombination. In classical recombination, a linear double stranded DNA with a synthetic DNA (s. DNA) sequence and a positive selection marker (+, Cm R ) flanked by homologous region 1 (HR1) and homologous region 2 (HR2) is transformed into the cell. Recombinants are selected by expression of the positive selection marker. By simultaneous double selection recombination, s. DNA containing double selection marker -2/+2 ( sacB-Cm R ) is integrated in place of the double selection marker -1/+1 ( rpsL - Kan R ) on the genome. Double selection for the gain of +2 and loss of -1 selects for simultaneous gain of s. DNA and loss of genomic sequence, and improves recombination at the target genomic locus. b. Colony PCR of clones from classical recombination and simultaneous double selection and recombination. c. All of the clones isolated by simultaneous double selection and recombination have s. DNA integrated at the target locus. The data show the mean of three independent experiments, the error bars represent the standard deviation (n=6). d . Both simultaneous double selection recombination (n = 8), and REXER 2 and REXER 4 (n = 296) result in the right combination of markers. A previously reported method integrating foreign DNA into B. subtilis , . A previously reported method replacing S. cerevisiae .

    Techniques Used: Selection, Sequencing, Marker, Transformation Assay, Expressing, Polymerase Chain Reaction, Clone Assay, Isolation, Standard Deviation

    35) Product Images from "Modifications and optimization of manual methods for polymerase chain reaction and 16S rRNA gene sequencing quality community DNA extraction from goat rumen digesta"

    Article Title: Modifications and optimization of manual methods for polymerase chain reaction and 16S rRNA gene sequencing quality community DNA extraction from goat rumen digesta

    Journal: Veterinary World

    doi: 10.14202/vetworld.2018.990-1000

    (a and b) Standard polymerase chain reaction (PCR) using community DNA extracted using modified methods. Standard PCR using (a) universal bacterial primers with community DNA extracted using Lane 1: 1 Kb ladder (New England Biolabs), Lane 2: Enzymatic method (EM)1, Lane 3: EC1, Lane 4: Enzymatic+Chemical+Physical method (ECPM)2, Lane 5: Chemical method (CM)1, Lane 6: CM4, Lane 7: CM2 methods, Lane 9: Genomic DNA of Bacillus subtilis as a positive control and Lane 10: blank (b) Specific targeted bacterial 16S rRNA gene primers and community DNA as follows. Lane 1: 100 bp ladder (New England Biolabs), Lane 2: Blank, Lane 3-4: CM4 and ECPM2 DNA with genus Bacteroides and Prevotella (418 bp), Lane 5-6: CM4 and ECPM2 DNA with Streptococcus bovis (869 bp), Lane 7-8: CM4 and ECPM2 DNA with Ruminococcus flavefaciens (835 bp), Lane 9-10: CM4 and ECPM2 DNA with Fibrobacter succinogenes (446 bp), and Lane 11-12: CM4 and ECPM2 DNA with Selenomonas ruminantium (513 bp), and Lane 13: blank.
    Figure Legend Snippet: (a and b) Standard polymerase chain reaction (PCR) using community DNA extracted using modified methods. Standard PCR using (a) universal bacterial primers with community DNA extracted using Lane 1: 1 Kb ladder (New England Biolabs), Lane 2: Enzymatic method (EM)1, Lane 3: EC1, Lane 4: Enzymatic+Chemical+Physical method (ECPM)2, Lane 5: Chemical method (CM)1, Lane 6: CM4, Lane 7: CM2 methods, Lane 9: Genomic DNA of Bacillus subtilis as a positive control and Lane 10: blank (b) Specific targeted bacterial 16S rRNA gene primers and community DNA as follows. Lane 1: 100 bp ladder (New England Biolabs), Lane 2: Blank, Lane 3-4: CM4 and ECPM2 DNA with genus Bacteroides and Prevotella (418 bp), Lane 5-6: CM4 and ECPM2 DNA with Streptococcus bovis (869 bp), Lane 7-8: CM4 and ECPM2 DNA with Ruminococcus flavefaciens (835 bp), Lane 9-10: CM4 and ECPM2 DNA with Fibrobacter succinogenes (446 bp), and Lane 11-12: CM4 and ECPM2 DNA with Selenomonas ruminantium (513 bp), and Lane 13: blank.

    Techniques Used: Polymerase Chain Reaction, Modification, Positive Control

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    New England Biolabs 1 kb ladder
    Linearized plasmid profiles of Edwardsiella ictaluri isolates. Plasmid DNA from E. ictaluri isolated from Channel Catfish or Zebrafish was digested with EcoRI or BstZ17I, respectively, and separated by 0.6% agarose gel electrophoresis using a <t>1-kb</t> DNA
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    Linearized plasmid profiles of Edwardsiella ictaluri isolates. Plasmid DNA from E. ictaluri isolated from Channel Catfish or Zebrafish was digested with EcoRI or BstZ17I, respectively, and separated by 0.6% agarose gel electrophoresis using a 1-kb DNA

    Journal: Journal of aquatic animal health

    Article Title: Edwardsiellosis Caused by Edwardsiella ictaluri in Laboratory Populations of Zebrafish Danio rerio

    doi: 10.1080/08997659.2013.782226

    Figure Lengend Snippet: Linearized plasmid profiles of Edwardsiella ictaluri isolates. Plasmid DNA from E. ictaluri isolated from Channel Catfish or Zebrafish was digested with EcoRI or BstZ17I, respectively, and separated by 0.6% agarose gel electrophoresis using a 1-kb DNA

    Article Snippet: Plasmids were digested with either EcoRI or BstZ17I and were separated by 0.6% agarose gel electrophoresis with 1-kb ladder (New England Biolabs) as the size standard.

    Techniques: Plasmid Preparation, Isolation, Agarose Gel Electrophoresis

    1% agarose gel electrophoresis of PCR products . PCR products derived from 24 PLUG primer sets were separated using a 1% agarose gel in TAE buffer. Lane numbers correspond to marker numbers indicated in Table 1. M: 2-Log DNA Ladder (New England BioLabs Inc., Ipswich, MA, USA).

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    Figure Lengend Snippet: 1% agarose gel electrophoresis of PCR products . PCR products derived from 24 PLUG primer sets were separated using a 1% agarose gel in TAE buffer. Lane numbers correspond to marker numbers indicated in Table 1. M: 2-Log DNA Ladder (New England BioLabs Inc., Ipswich, MA, USA).

    Article Snippet: Band sizes were estimated against a '2-Log DNA Ladder' (New England BioLabs Inc., Ipswich, MA, USA).

    Techniques: Agarose Gel Electrophoresis, Polymerase Chain Reaction, Derivative Assay, Marker

    Subcellular localization of PRP1 through endogenous tagging. (A) Schematic representation of generating C-terminal endogenously YFP-tagged gPRP1-YFP parasites by single homologous recombination into the RHΔ ku80 parent line. (B) PCR validation of the gPRP1-YFP genotype using the primer pair shown in panel A. Lane M contains 1-kb DNA ladder (New England Biolabs). (C) Live imaging of gPRP1-YFP parasites under intracellular and extracellular conditions as indicated. PC, phase contrast. (D) Live imaging of gPRP1-YFP parasites cotransfected with markers for the IMC (IMC1-mCherry), rhoptries (TLN1-mCherry), and micronemes (MIC8-mCherry). (E) Representative images of intracellular gPRP1-YFP parasites fixed using either 100% methanol (MetOH) or 4% paraformaldehyde (PFA) stained with anti-PRP1 (αPRP1) and anti-GFP (αGFP) antisera as indicated. Note that PFA fixation destroys the costaining of GFP and PRP1 and thus destroys the PRP1 epitope(s) recognized by the specific antiserum.

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    Figure Lengend Snippet: Subcellular localization of PRP1 through endogenous tagging. (A) Schematic representation of generating C-terminal endogenously YFP-tagged gPRP1-YFP parasites by single homologous recombination into the RHΔ ku80 parent line. (B) PCR validation of the gPRP1-YFP genotype using the primer pair shown in panel A. Lane M contains 1-kb DNA ladder (New England Biolabs). (C) Live imaging of gPRP1-YFP parasites under intracellular and extracellular conditions as indicated. PC, phase contrast. (D) Live imaging of gPRP1-YFP parasites cotransfected with markers for the IMC (IMC1-mCherry), rhoptries (TLN1-mCherry), and micronemes (MIC8-mCherry). (E) Representative images of intracellular gPRP1-YFP parasites fixed using either 100% methanol (MetOH) or 4% paraformaldehyde (PFA) stained with anti-PRP1 (αPRP1) and anti-GFP (αGFP) antisera as indicated. Note that PFA fixation destroys the costaining of GFP and PRP1 and thus destroys the PRP1 epitope(s) recognized by the specific antiserum.

    Article Snippet: M represents 1-kb DNA ladder (NEB).

    Techniques: Homologous Recombination, Polymerase Chain Reaction, Imaging, Staining

    Gel electrophoresis confirming stability of plasmid after ultrasound exposures. Lane 1) pDNA; lane 2) sonicated pDNA; lane 3) lipoplex; lane 4) sonicated lipoplex; lane 5) emulsified lipoplex released by ultrasound (3.5 MHz, mechanical index (MI) = 2.5, 10 Hz pulse repetition frequency (PRF), 30 cycles); lane 6) 1 kb linear DNA ladder marker. The mass of pDNA was equivalent in all lanes.

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    Figure Lengend Snippet: Gel electrophoresis confirming stability of plasmid after ultrasound exposures. Lane 1) pDNA; lane 2) sonicated pDNA; lane 3) lipoplex; lane 4) sonicated lipoplex; lane 5) emulsified lipoplex released by ultrasound (3.5 MHz, mechanical index (MI) = 2.5, 10 Hz pulse repetition frequency (PRF), 30 cycles); lane 6) 1 kb linear DNA ladder marker. The mass of pDNA was equivalent in all lanes.

    Article Snippet: Control samples of non-sonicated plasmid and lipoplex as well as a 1 kb linear DNA ladder (N3232S, New England BioLabs, Ipswich, MA, USA) were also run on the gel.

    Techniques: Nucleic Acid Electrophoresis, Plasmid Preparation, Sonication, Marker