exonuclease iii digestion Search Results


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  • 99
    New England Biolabs exonuclease iii digestion
    HCc3 promotes ligase-mediated DNA concatenation. All images are presented as negative images. ( a ) Concatenation of 125-bp (left panel) and 2.8-kb (right panel) DNA fragments to a higher level in the presence of HCc3 at respective dimer/bp ratios. The linearity of the resulting products was confirmed by <t>Exo</t> <t>III</t> digestion (marked with ‘+/−’ signs). The 125-bp product was resolved on a 8% non-denaturing polyacrylamide gel, and the 2.8-kb product on a 1% TAE agarose gel. ( b ) Variations in intensity of DNA concatenation in the presence of HCc3 at different dimer/bp ratios. The marker bands indicate DNA sizes from 3 to 12 kb at 1-kb increments.
    Exonuclease Iii Digestion, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 64 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Thermo Fisher exonuclease iii
    Figure 2. <t>PCR</t> checkpoint after clonal enrichment. ( A ) An agarose gel stained with ethidium bromide with inverted grayscale showing the PCR products of the clonal enrichment of group 2G3 and a negative control (mock enrichment). Two main products were amplified, one covering the whole VH region (VH-JH) and the other up to the CDRH3 (IGHV-CDRH3), of approximately 350 and 300 base pair, respectively. <t>Three</t> different amplification cycles were used (13X, 15X, and 18X) to determine the amplification threshold required for further processing. In the enriched sample, both products (IGHV-IGHJ and IGHV-CDRH3) are present, while they are slightly visible in 18X of the mock control. This difference demonstrates a selection for VH sequences containing the desired CDRH3. ( B ) PCR products of the ten enriched samples and their respective mock controls shown in separate agarose gels stained with ethidium bromide with an inverted grayscale. For each group, the enriched sample (left) and mock control (right) amplifications were performed with the indicated amplification cycles and annealing temperatures. The PCR products from the enriched samples shown here were used for cloning and sequencing.
    Exonuclease Iii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 295 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Promega exonuclease iii digestion
    Repression of <t>HIF-1β</t> promoter activity by IFN-γ. A , Cloned HIF-1β promoter and series of truncated promoter-luciferase reporter constructs generated using exonuclease <t>III</t> digestion. Relative positions of each clone and major TSSs
    Exonuclease Iii Digestion, supplied by Promega, used in various techniques. Bioz Stars score: 88/100, based on 98 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    TaKaRa exonuclease iii
    The MALDI-TOF MS spectra for genotyping of SNP in apoE gene by using exonuclease <t>III/nuclease</t> S1/PNA systems. ( A ) Analysis of dsDNA from apoE 4 (250 bp) at codon 112 using PNA (S112G) . Reaction conditions: [ apoE <t>DNA]</t> = 3 µM,
    Exonuclease Iii, supplied by TaKaRa, used in various techniques. Bioz Stars score: 93/100, based on 174 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Millipore exonuclease iii digestion
    The MALDI-TOF MS spectra for genotyping of SNP in apoE gene by using exonuclease <t>III/nuclease</t> S1/PNA systems. ( A ) Analysis of dsDNA from apoE 4 (250 bp) at codon 112 using PNA (S112G) . Reaction conditions: [ apoE <t>DNA]</t> = 3 µM,
    Exonuclease Iii Digestion, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    TaKaRa exonuclease iii digestion
    The distal region of the IL-6 promoter contains functional C/EBP motifs A. IL-6 promoter Luciferase activity in PC3 cells transfected with the <t>pGL2-IL-6</t> deletion mutants shown on the left and pRV with and without treatment with rGal-3BP (2.5 µg/ml) for 24 hours. The data represent the mean (±SD) ratio Firefly Luciferase/Renilla Luciferase and are representative of one among <t>three</t> separate experiments each performed in triplicate samples. B. Sequencing analysis of the region of the IL-6 promoter extending from position −1586 to −1255 showing the presence of 3 C/EBP domains. C. EMSA of nuclear extracts from PC3 cells treated with rGAL-3BP (5 µg/ml) using biotinylated oligonucleotides in the presence of 10× fold excess of non-biotinylated oligonucleotides when indicated.
    Exonuclease Iii Digestion, supplied by TaKaRa, used in various techniques. Bioz Stars score: 96/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Promega exonuclease iii solution
    The distal region of the IL-6 promoter contains functional C/EBP motifs A. IL-6 promoter Luciferase activity in PC3 cells transfected with the <t>pGL2-IL-6</t> deletion mutants shown on the left and pRV with and without treatment with rGal-3BP (2.5 µg/ml) for 24 hours. The data represent the mean (±SD) ratio Firefly Luciferase/Renilla Luciferase and are representative of one among <t>three</t> separate experiments each performed in triplicate samples. B. Sequencing analysis of the region of the IL-6 promoter extending from position −1586 to −1255 showing the presence of 3 C/EBP domains. C. EMSA of nuclear extracts from PC3 cells treated with rGAL-3BP (5 µg/ml) using biotinylated oligonucleotides in the presence of 10× fold excess of non-biotinylated oligonucleotides when indicated.
    Exonuclease Iii Solution, supplied by Promega, used in various techniques. Bioz Stars score: 93/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Thermo Fisher lambda exonuclease iii
    The distal region of the IL-6 promoter contains functional C/EBP motifs A. IL-6 promoter Luciferase activity in PC3 cells transfected with the <t>pGL2-IL-6</t> deletion mutants shown on the left and pRV with and without treatment with rGal-3BP (2.5 µg/ml) for 24 hours. The data represent the mean (±SD) ratio Firefly Luciferase/Renilla Luciferase and are representative of one among <t>three</t> separate experiments each performed in triplicate samples. B. Sequencing analysis of the region of the IL-6 promoter extending from position −1586 to −1255 showing the presence of 3 C/EBP domains. C. EMSA of nuclear extracts from PC3 cells treated with rGAL-3BP (5 µg/ml) using biotinylated oligonucleotides in the presence of 10× fold excess of non-biotinylated oligonucleotides when indicated.
    Lambda Exonuclease Iii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 95/100, based on 21 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Thermo Fisher exonucleases iii
    The distal region of the IL-6 promoter contains functional C/EBP motifs A. IL-6 promoter Luciferase activity in PC3 cells transfected with the <t>pGL2-IL-6</t> deletion mutants shown on the left and pRV with and without treatment with rGal-3BP (2.5 µg/ml) for 24 hours. The data represent the mean (±SD) ratio Firefly Luciferase/Renilla Luciferase and are representative of one among <t>three</t> separate experiments each performed in triplicate samples. B. Sequencing analysis of the region of the IL-6 promoter extending from position −1586 to −1255 showing the presence of 3 C/EBP domains. C. EMSA of nuclear extracts from PC3 cells treated with rGAL-3BP (5 µg/ml) using biotinylated oligonucleotides in the presence of 10× fold excess of non-biotinylated oligonucleotides when indicated.
    Exonucleases Iii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 88/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore dnase i protease inhibitor cocktail set iii
    The distal region of the IL-6 promoter contains functional C/EBP motifs A. IL-6 promoter Luciferase activity in PC3 cells transfected with the <t>pGL2-IL-6</t> deletion mutants shown on the left and pRV with and without treatment with rGal-3BP (2.5 µg/ml) for 24 hours. The data represent the mean (±SD) ratio Firefly Luciferase/Renilla Luciferase and are representative of one among <t>three</t> separate experiments each performed in triplicate samples. B. Sequencing analysis of the region of the IL-6 promoter extending from position −1586 to −1255 showing the presence of 3 C/EBP domains. C. EMSA of nuclear extracts from PC3 cells treated with rGAL-3BP (5 µg/ml) using biotinylated oligonucleotides in the presence of 10× fold excess of non-biotinylated oligonucleotides when indicated.
    Dnase I Protease Inhibitor Cocktail Set Iii, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore type iii collagenase dnase
    The distal region of the IL-6 promoter contains functional C/EBP motifs A. IL-6 promoter Luciferase activity in PC3 cells transfected with the <t>pGL2-IL-6</t> deletion mutants shown on the left and pRV with and without treatment with rGal-3BP (2.5 µg/ml) for 24 hours. The data represent the mean (±SD) ratio Firefly Luciferase/Renilla Luciferase and are representative of one among <t>three</t> separate experiments each performed in triplicate samples. B. Sequencing analysis of the region of the IL-6 promoter extending from position −1586 to −1255 showing the presence of 3 C/EBP domains. C. EMSA of nuclear extracts from PC3 cells treated with rGAL-3BP (5 µg/ml) using biotinylated oligonucleotides in the presence of 10× fold excess of non-biotinylated oligonucleotides when indicated.
    Type Iii Collagenase Dnase, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    New England Biolabs digestion mixture
    The distal region of the IL-6 promoter contains functional C/EBP motifs A. IL-6 promoter Luciferase activity in PC3 cells transfected with the <t>pGL2-IL-6</t> deletion mutants shown on the left and pRV with and without treatment with rGal-3BP (2.5 µg/ml) for 24 hours. The data represent the mean (±SD) ratio Firefly Luciferase/Renilla Luciferase and are representative of one among <t>three</t> separate experiments each performed in triplicate samples. B. Sequencing analysis of the region of the IL-6 promoter extending from position −1586 to −1255 showing the presence of 3 C/EBP domains. C. EMSA of nuclear extracts from PC3 cells treated with rGAL-3BP (5 µg/ml) using biotinylated oligonucleotides in the presence of 10× fold excess of non-biotinylated oligonucleotides when indicated.
    Digestion Mixture, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Thermo Fisher exo iii s1n digestion
    The distal region of the IL-6 promoter contains functional C/EBP motifs A. IL-6 promoter Luciferase activity in PC3 cells transfected with the <t>pGL2-IL-6</t> deletion mutants shown on the left and pRV with and without treatment with rGal-3BP (2.5 µg/ml) for 24 hours. The data represent the mean (±SD) ratio Firefly Luciferase/Renilla Luciferase and are representative of one among <t>three</t> separate experiments each performed in triplicate samples. B. Sequencing analysis of the region of the IL-6 promoter extending from position −1586 to −1255 showing the presence of 3 C/EBP domains. C. EMSA of nuclear extracts from PC3 cells treated with rGAL-3BP (5 µg/ml) using biotinylated oligonucleotides in the presence of 10× fold excess of non-biotinylated oligonucleotides when indicated.
    Exo Iii S1n Digestion, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Promega iii si nuclease digestion
    The distal region of the IL-6 promoter contains functional C/EBP motifs A. IL-6 promoter Luciferase activity in PC3 cells transfected with the <t>pGL2-IL-6</t> deletion mutants shown on the left and pRV with and without treatment with rGal-3BP (2.5 µg/ml) for 24 hours. The data represent the mean (±SD) ratio Firefly Luciferase/Renilla Luciferase and are representative of one among <t>three</t> separate experiments each performed in triplicate samples. B. Sequencing analysis of the region of the IL-6 promoter extending from position −1586 to −1255 showing the presence of 3 C/EBP domains. C. EMSA of nuclear extracts from PC3 cells treated with rGAL-3BP (5 µg/ml) using biotinylated oligonucleotides in the presence of 10× fold excess of non-biotinylated oligonucleotides when indicated.
    Iii Si Nuclease Digestion, supplied by Promega, used in various techniques. Bioz Stars score: 85/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    HCc3 promotes ligase-mediated DNA concatenation. All images are presented as negative images. ( a ) Concatenation of 125-bp (left panel) and 2.8-kb (right panel) DNA fragments to a higher level in the presence of HCc3 at respective dimer/bp ratios. The linearity of the resulting products was confirmed by Exo III digestion (marked with ‘+/−’ signs). The 125-bp product was resolved on a 8% non-denaturing polyacrylamide gel, and the 2.8-kb product on a 1% TAE agarose gel. ( b ) Variations in intensity of DNA concatenation in the presence of HCc3 at different dimer/bp ratios. The marker bands indicate DNA sizes from 3 to 12 kb at 1-kb increments.

    Journal: Nucleic Acids Research

    Article Title: Concentration-dependent organization of DNA by the dinoflagellate histone-like protein HCc3

    doi: 10.1093/nar/gkm165

    Figure Lengend Snippet: HCc3 promotes ligase-mediated DNA concatenation. All images are presented as negative images. ( a ) Concatenation of 125-bp (left panel) and 2.8-kb (right panel) DNA fragments to a higher level in the presence of HCc3 at respective dimer/bp ratios. The linearity of the resulting products was confirmed by Exo III digestion (marked with ‘+/−’ signs). The 125-bp product was resolved on a 8% non-denaturing polyacrylamide gel, and the 2.8-kb product on a 1% TAE agarose gel. ( b ) Variations in intensity of DNA concatenation in the presence of HCc3 at different dimer/bp ratios. The marker bands indicate DNA sizes from 3 to 12 kb at 1-kb increments.

    Article Snippet: If samples were to be treated with Exonuclease III (Exo III) digestion, an equal volume of 2× digestion mixture (2× NEBuffer 1 and 1 U/μl Exonuclease III; New England Biolab) was added to the reaction and the digestion occurred at 37°C for 1 h, prior to the addition of stop solution.

    Techniques: Agarose Gel Electrophoresis, Marker

    DNA repair pathways implicated in 5-FU-mediated cell killing. The model is supported by the following observations: (i) a massive amount of uracil is incorporated into DNA, but the ung1 yeast are much less sensitive to 5-FU than the wild-type strain indicating that uracilated DNA is not the mediator of 5-FU toxicity; (ii) the apn1apn2ntg1ntg2 strain that is entirely defective in processing abasic sites by a BER mechanism is more sensitive to 5-FU, indicating that intact abasic sites (or repair products derived from abasic sites) have inherent toxicity; and (iii) the rad27 and apn1rad27 yeast strains show protection against 5-FU toxicity, suggesting the presence of a toxic repair intermediate downstream of the Rad27 flap endonuclease reaction. Several backup pathways for repair of abasic sites and 5′dRp groups are indicated. The lower path involving Apn2 and other BER enzymes is important in the absence of Apn1 and accounts for the efficient removal of abasic sites in the apn1 strain. NER and HR pathways are likely to be important with the apn1apn2ntg1ntg2 and rad27 knockout strains. Consistent with this, yeast deficient in both BER and NER are not viable.

    Journal: Nucleic Acids Research

    Article Title: Linking uracil base excision repair and 5-fluorouracil toxicity in yeast

    doi: 10.1093/nar/gkj430

    Figure Lengend Snippet: DNA repair pathways implicated in 5-FU-mediated cell killing. The model is supported by the following observations: (i) a massive amount of uracil is incorporated into DNA, but the ung1 yeast are much less sensitive to 5-FU than the wild-type strain indicating that uracilated DNA is not the mediator of 5-FU toxicity; (ii) the apn1apn2ntg1ntg2 strain that is entirely defective in processing abasic sites by a BER mechanism is more sensitive to 5-FU, indicating that intact abasic sites (or repair products derived from abasic sites) have inherent toxicity; and (iii) the rad27 and apn1rad27 yeast strains show protection against 5-FU toxicity, suggesting the presence of a toxic repair intermediate downstream of the Rad27 flap endonuclease reaction. Several backup pathways for repair of abasic sites and 5′dRp groups are indicated. The lower path involving Apn2 and other BER enzymes is important in the absence of Apn1 and accounts for the efficient removal of abasic sites in the apn1 strain. NER and HR pathways are likely to be important with the apn1apn2ntg1ntg2 and rad27 knockout strains. Consistent with this, yeast deficient in both BER and NER are not viable.

    Article Snippet: Briefly, 4 µg of each DNA sample was digested with E.coli exonuclease III (145 U; New England Biolabs) for 1 min at 37°C, 100 mM putrescine at 37°C for 30 min (Acros Organics), both exonuclease III and putrescine, or left undigested.

    Techniques: Derivative Assay, Knock-Out

    Figure 2. PCR checkpoint after clonal enrichment. ( A ) An agarose gel stained with ethidium bromide with inverted grayscale showing the PCR products of the clonal enrichment of group 2G3 and a negative control (mock enrichment). Two main products were amplified, one covering the whole VH region (VH-JH) and the other up to the CDRH3 (IGHV-CDRH3), of approximately 350 and 300 base pair, respectively. Three different amplification cycles were used (13X, 15X, and 18X) to determine the amplification threshold required for further processing. In the enriched sample, both products (IGHV-IGHJ and IGHV-CDRH3) are present, while they are slightly visible in 18X of the mock control. This difference demonstrates a selection for VH sequences containing the desired CDRH3. ( B ) PCR products of the ten enriched samples and their respective mock controls shown in separate agarose gels stained with ethidium bromide with an inverted grayscale. For each group, the enriched sample (left) and mock control (right) amplifications were performed with the indicated amplification cycles and annealing temperatures. The PCR products from the enriched samples shown here were used for cloning and sequencing.

    Journal: mAbs

    Article Title: Hybridization-based antibody cDNA recovery for the production of recombinant antibodies identified by repertoire sequencing

    doi: 10.4161/mabs.27435

    Figure Lengend Snippet: Figure 2. PCR checkpoint after clonal enrichment. ( A ) An agarose gel stained with ethidium bromide with inverted grayscale showing the PCR products of the clonal enrichment of group 2G3 and a negative control (mock enrichment). Two main products were amplified, one covering the whole VH region (VH-JH) and the other up to the CDRH3 (IGHV-CDRH3), of approximately 350 and 300 base pair, respectively. Three different amplification cycles were used (13X, 15X, and 18X) to determine the amplification threshold required for further processing. In the enriched sample, both products (IGHV-IGHJ and IGHV-CDRH3) are present, while they are slightly visible in 18X of the mock control. This difference demonstrates a selection for VH sequences containing the desired CDRH3. ( B ) PCR products of the ten enriched samples and their respective mock controls shown in separate agarose gels stained with ethidium bromide with an inverted grayscale. For each group, the enriched sample (left) and mock control (right) amplifications were performed with the indicated amplification cycles and annealing temperatures. The PCR products from the enriched samples shown here were used for cloning and sequencing.

    Article Snippet: The new PCR products were digested with Exonuclease III, and the pVAJO-CHG1 with EcoRI and NheI (Fermentas).

    Techniques: Polymerase Chain Reaction, Agarose Gel Electrophoresis, Staining, Negative Control, Amplification, Selection, Clone Assay, Sequencing

    Figure 3. Enrichment assessment through single sequence analysis. ( A ) Representation of the concentrations of each of the eight clonal groups that were successfully recovered, before (gray) and after (yellow) the enrichment. Concentrations before enrichment were determined in silico by the analysis of the massive sequencing results; the number of sequences considered for 2G1–2G5 was 46 062, and 36 393 for 3G1–3G4. Concentrations after enrichment were determined through PCR amplification of the desired IGHV-CDRH3 fragment from cloned enrichment products; N indicates the total number of clones evaluated in each case. The circle representing the total (blue) is used as a reference of 100% of the area. ( B ) CDRH3 conservation after enrichment determined through Sanger sequencing. A few clones for each clonal group were sequenced (N) and compared with their respective target sequence, showing that most of the sequences obtained had no or little variation in the CDRH3. ( C ) HEL-binding ELISA for the three positive combinations (VH/VL) of the 32 tested. 2G3/HH10, 3G1/HH5 and 3G1/F10.6.6 showed HEL-binding capacity, corresponding to two VH of the eight tested. HH10/HH10 served as positive control, and no-DNA transfectant was used as negative control. The y-axis shows the ratio between the OD 490 from the HEL-binding ELISA and the OD 490 from the IgG detection ELISA of the same sample.

    Journal: mAbs

    Article Title: Hybridization-based antibody cDNA recovery for the production of recombinant antibodies identified by repertoire sequencing

    doi: 10.4161/mabs.27435

    Figure Lengend Snippet: Figure 3. Enrichment assessment through single sequence analysis. ( A ) Representation of the concentrations of each of the eight clonal groups that were successfully recovered, before (gray) and after (yellow) the enrichment. Concentrations before enrichment were determined in silico by the analysis of the massive sequencing results; the number of sequences considered for 2G1–2G5 was 46 062, and 36 393 for 3G1–3G4. Concentrations after enrichment were determined through PCR amplification of the desired IGHV-CDRH3 fragment from cloned enrichment products; N indicates the total number of clones evaluated in each case. The circle representing the total (blue) is used as a reference of 100% of the area. ( B ) CDRH3 conservation after enrichment determined through Sanger sequencing. A few clones for each clonal group were sequenced (N) and compared with their respective target sequence, showing that most of the sequences obtained had no or little variation in the CDRH3. ( C ) HEL-binding ELISA for the three positive combinations (VH/VL) of the 32 tested. 2G3/HH10, 3G1/HH5 and 3G1/F10.6.6 showed HEL-binding capacity, corresponding to two VH of the eight tested. HH10/HH10 served as positive control, and no-DNA transfectant was used as negative control. The y-axis shows the ratio between the OD 490 from the HEL-binding ELISA and the OD 490 from the IgG detection ELISA of the same sample.

    Article Snippet: The new PCR products were digested with Exonuclease III, and the pVAJO-CHG1 with EcoRI and NheI (Fermentas).

    Techniques: Sequencing, In Silico, Polymerase Chain Reaction, Amplification, Clone Assay, Binding Assay, Enzyme-linked Immunosorbent Assay, Positive Control, Transfection, Negative Control

    Repression of HIF-1β promoter activity by IFN-γ. A , Cloned HIF-1β promoter and series of truncated promoter-luciferase reporter constructs generated using exonuclease III digestion. Relative positions of each clone and major TSSs

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    Article Title: IFN-? Attenuates Hypoxia-Inducible Factor (HIF) Activity in Intestinal Epithelial Cells through Transcriptional Repression of HIF-1?

    doi: 10.4049/jimmunol.1001442

    Figure Lengend Snippet: Repression of HIF-1β promoter activity by IFN-γ. A , Cloned HIF-1β promoter and series of truncated promoter-luciferase reporter constructs generated using exonuclease III digestion. Relative positions of each clone and major TSSs

    Article Snippet: Sequential truncations of the cloned HIF-1β promoter sequence were generated by exonuclease III digestion (Erase-a-Base; Promega).

    Techniques: Activity Assay, Clone Assay, Luciferase, Construct, Generated

    Schema of ReDFISH technique. ( a ) ReDFISH of a chromosome that has replicated fully in the presence of BrdUrd/dC. Newly synthesized DNA incorporating BrdUrd/dC (horizontal stripes) is removed after nicking the DNA with Hoechst 33258 plus UV and digesting nicked DNA with exonuclease III, leaving only the parental strands. The G-rich telomeric strand is the template for lagging strand synthesis and anneals to a Cy3-conjugated C-rich telomeric probe, whereas the C-rich telomeric strand is the template for leading strand synthesis and anneals to an FITC-conjugated G-rich telomeric probe. This pattern defines which telomeric strands replicated at the time of BrdUrd/BrdC labeling. ( b ) ReDFISH of a partially replicated chromosome. In this example, only the p-arm telomere of the X chromosome was replicating during the 1-h BrdUrd/dC pulse; the q arm was not replicating. As a consequence, only the parental strands are available for hybridization in the p arm (schema, p arms), whereas both strands of unreplicated q-arm DNA survive digestion and hybridize to both probes (schema, q arms).

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Asynchronous replication timing of telomeres at opposite arms of mammalian chromosomes

    doi: 10.1073/pnas.0404106101

    Figure Lengend Snippet: Schema of ReDFISH technique. ( a ) ReDFISH of a chromosome that has replicated fully in the presence of BrdUrd/dC. Newly synthesized DNA incorporating BrdUrd/dC (horizontal stripes) is removed after nicking the DNA with Hoechst 33258 plus UV and digesting nicked DNA with exonuclease III, leaving only the parental strands. The G-rich telomeric strand is the template for lagging strand synthesis and anneals to a Cy3-conjugated C-rich telomeric probe, whereas the C-rich telomeric strand is the template for leading strand synthesis and anneals to an FITC-conjugated G-rich telomeric probe. This pattern defines which telomeric strands replicated at the time of BrdUrd/BrdC labeling. ( b ) ReDFISH of a partially replicated chromosome. In this example, only the p-arm telomere of the X chromosome was replicating during the 1-h BrdUrd/dC pulse; the q arm was not replicating. As a consequence, only the parental strands are available for hybridization in the p arm (schema, p arms), whereas both strands of unreplicated q-arm DNA survive digestion and hybridize to both probes (schema, q arms).

    Article Snippet: The nicked BrdUrd/dC-substituted DNA was digested with 3 units/μl exonuclease III (Promega) in 50 mM Tris·HCl (pH 8.0), 5 mM MgCl2 , and 5 mM DTT for 10 min at room temperature.

    Techniques: Synthesized, Labeling, Hybridization

    The MALDI-TOF MS spectra for genotyping of SNP in apoE gene by using exonuclease III/nuclease S1/PNA systems. ( A ) Analysis of dsDNA from apoE 4 (250 bp) at codon 112 using PNA (S112G) . Reaction conditions: [ apoE DNA] = 3 µM,

    Journal: Nucleic Acids Research

    Article Title: Straightforward detection of SNPs in double-stranded DNA by using exonuclease III/nuclease S1/PNA system

    doi: 10.1093/nar/gnh039

    Figure Lengend Snippet: The MALDI-TOF MS spectra for genotyping of SNP in apoE gene by using exonuclease III/nuclease S1/PNA systems. ( A ) Analysis of dsDNA from apoE 4 (250 bp) at codon 112 using PNA (S112G) . Reaction conditions: [ apoE DNA] = 3 µM,

    Article Snippet: The substrate DNA was digested by exonuclease III ( TaKaRa Bio Inc., Tokyo) in 1× exonuclease III buffer at pH 8.0 (50 mM Tris–HCl, 5 mM MgCl2 , 10 mM 2-mercaptoethanol) and 37°C for 5 or 10 min. Then, 1× nuclease S1 buffer pH 4.6 (30 mM sodium acetate, 1 mM zinc acetate, 5% (v/v) glycerol) and PNA additives were added (the volume ratio of the nuclease S1 buffer to the exonuclease III buffer was 1.5:1.0).

    Techniques: Mass Spectrometry

    The distal region of the IL-6 promoter contains functional C/EBP motifs A. IL-6 promoter Luciferase activity in PC3 cells transfected with the pGL2-IL-6 deletion mutants shown on the left and pRV with and without treatment with rGal-3BP (2.5 µg/ml) for 24 hours. The data represent the mean (±SD) ratio Firefly Luciferase/Renilla Luciferase and are representative of one among three separate experiments each performed in triplicate samples. B. Sequencing analysis of the region of the IL-6 promoter extending from position −1586 to −1255 showing the presence of 3 C/EBP domains. C. EMSA of nuclear extracts from PC3 cells treated with rGAL-3BP (5 µg/ml) using biotinylated oligonucleotides in the presence of 10× fold excess of non-biotinylated oligonucleotides when indicated.

    Journal: Cancer research

    Article Title: A GALECTIN-3-DEPENDENT PATHWAY UPREGULATES INTERLEUKIN-6 IN THE MICROENVIRONMENT OF HUMAN NEUROBLASTOMA

    doi: 10.1158/0008-5472.CAN-11-2165

    Figure Lengend Snippet: The distal region of the IL-6 promoter contains functional C/EBP motifs A. IL-6 promoter Luciferase activity in PC3 cells transfected with the pGL2-IL-6 deletion mutants shown on the left and pRV with and without treatment with rGal-3BP (2.5 µg/ml) for 24 hours. The data represent the mean (±SD) ratio Firefly Luciferase/Renilla Luciferase and are representative of one among three separate experiments each performed in triplicate samples. B. Sequencing analysis of the region of the IL-6 promoter extending from position −1586 to −1255 showing the presence of 3 C/EBP domains. C. EMSA of nuclear extracts from PC3 cells treated with rGAL-3BP (5 µg/ml) using biotinylated oligonucleotides in the presence of 10× fold excess of non-biotinylated oligonucleotides when indicated.

    Article Snippet: IL-6 promoter deletion mutants in the pGL2-IL-6-Luc construct were generated either by Exonuclease III digestion using the Deletion Kit for kilo sequencing (Takara) in accordance with the instructions of the manufacturer (deletion −1041) or by restriction endonuclease digestion with KpnI and NheI (deletion mutant −212) followed by Klenow fragment reaction at 37°C for 15 minutes.

    Techniques: Functional Assay, Luciferase, Activity Assay, Transfection, Sequencing