bstui  (New England Biolabs)


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    Name:
    BstUI
    Description:
    BstUI 5 000 units
    Catalog Number:
    r0518l
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    257
    Category:
    Restriction Enzymes
    Size:
    5 000 units
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    New England Biolabs bstui
    BstUI
    BstUI 5 000 units
    https://www.bioz.com/result/bstui/product/New England Biolabs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    bstui - by Bioz Stars, 2021-03
    94/100 stars

    Images

    1) Product Images from "Lack of HLA class II antigen expression in microsatellite unstable colorectal carcinomas is caused by mutations in HLA class II regulatory genes"

    Article Title: Lack of HLA class II antigen expression in microsatellite unstable colorectal carcinomas is caused by mutations in HLA class II regulatory genes

    Journal: International journal of cancer. Journal international du cancer

    doi: 10.1002/ijc.25106

    Analysis of CIITA promoter IV methylation in MSI-H CRC lesions (A) and cell lines (B). Two regions (GM1 and GM2) of the promoter were analyzed for each sample. Methylation was analyzed by enzymatic digest with BstUI following bisulfite treatment and PCR
    Figure Legend Snippet: Analysis of CIITA promoter IV methylation in MSI-H CRC lesions (A) and cell lines (B). Two regions (GM1 and GM2) of the promoter were analyzed for each sample. Methylation was analyzed by enzymatic digest with BstUI following bisulfite treatment and PCR

    Techniques Used: Methylation, Polymerase Chain Reaction

    2) Product Images from "Recurrent mutations in a SERPINC1 hotspot associate with venous thrombosis without apparent antithrombin deficiency"

    Article Title: Recurrent mutations in a SERPINC1 hotspot associate with venous thrombosis without apparent antithrombin deficiency

    Journal: Oncotarget

    doi: 10.18632/oncotarget.21365

    Identification of a mutational hotspot in SERPINC1 (A) Electropherogram of samples carrying all mutations affecting antithrombin Arg294 and Val295 residues. (B) Restriction fragments generated with BstUI on the PCR product of SERPINC1 exon 5 using the primers described in Materials and Methods. (C) Localization in the structure of antithrombin of affected residues in the native, activated, and latent configurations. The central A sheet is colored in blue, the reactive center loop in red, and s1B in green. Model building was performed by using SWISS-MODEL and the Swiss-PdbViewer programs [Guex N, Peitsch MC. SWISS-MODEL and the Swiss-PdbViewer: an environment for comparative protein modelling. Electrophoresis 1997; 18: 2714–23] ( http://www.expasy.ch/spdbv ).
    Figure Legend Snippet: Identification of a mutational hotspot in SERPINC1 (A) Electropherogram of samples carrying all mutations affecting antithrombin Arg294 and Val295 residues. (B) Restriction fragments generated with BstUI on the PCR product of SERPINC1 exon 5 using the primers described in Materials and Methods. (C) Localization in the structure of antithrombin of affected residues in the native, activated, and latent configurations. The central A sheet is colored in blue, the reactive center loop in red, and s1B in green. Model building was performed by using SWISS-MODEL and the Swiss-PdbViewer programs [Guex N, Peitsch MC. SWISS-MODEL and the Swiss-PdbViewer: an environment for comparative protein modelling. Electrophoresis 1997; 18: 2714–23] ( http://www.expasy.ch/spdbv ).

    Techniques Used: Generated, Polymerase Chain Reaction, Electrophoresis

    3) Product Images from "Evidence for Transgenerational Transmission of Epigenetic Tumor Susceptibility in Drosophila"

    Article Title: Evidence for Transgenerational Transmission of Epigenetic Tumor Susceptibility in Drosophila

    Journal: PLoS Genetics

    doi: 10.1371/journal.pgen.0030151

    Increased Methylation of ftz Regulatory Region in Kr +/− Animals and Its Inheritance (A) Schematic representation of the ftz-lacZ reporter, showing the minimal ftz 5′ regulatory region previously shown to be sufficient to drive expression of a ftz-lacZ reporter transgene in ftz patterns [ 30 ]. Arrows above and below the horizontal line represent PCR primers used to amplify a 778-bp fragment, encompassing the 620-bp minimal ftz enhancer (see Figure S2 for sequence). Bent arrow indicates the start of the lacZ sequence. Vertical bars above and below the line represent positions of recognition sequences for restriction enzymes BstUI (CGCG) and HaeIII (GGCC), respectively. (B–E) Time courses of restriction digests of genomic DNA with enzymes sensitive (BstUI) or insensitive (HaeIII) to methylated DNA are shown as agarose gel pictures and quantifications. Genomic DNA was isolated from Kr 1 /CyO ftz-lacZ and +/CyO ftz-lacZ (wild-type control) adult flies (B, C), or from the F1 progeny flies of Kr 1 /CyO ftz-lacZ males crossed to hop Tum-l/+ females or wild-type females (D, E), and digested with the indicated enzymes for the indicated times (minutes). Digested DNA was amplified with PCR primers shown in (A) and run on an agarose gel. Note that the genomic DNA from Kr +/ − flies or from the F1 progeny of hop Tum-l/+ females and Kr 1 /CyO ftz-lacZ males is more resistant to BstUI digestion than the controls. (F, G) Digested genomic DNA purified from embryos was immunoprecipitated by antibodies to methylated cytosine and amplified by PCR primers shown in (A). Embryos derived from wild-type or Kr +/ − parents (F), or from Kr 1 /ftz-lacZ males crossed to wild-type (top) or Tum-l/+ (bottom) females (G) were used for DNA isolation. Note the presence of higher levels of 5-meC in the ftz-lacZ promoter in Kr +/ − embryos or in those from Tum-l/+ females crossed to Kr 1 /ftz-lacZ males.
    Figure Legend Snippet: Increased Methylation of ftz Regulatory Region in Kr +/− Animals and Its Inheritance (A) Schematic representation of the ftz-lacZ reporter, showing the minimal ftz 5′ regulatory region previously shown to be sufficient to drive expression of a ftz-lacZ reporter transgene in ftz patterns [ 30 ]. Arrows above and below the horizontal line represent PCR primers used to amplify a 778-bp fragment, encompassing the 620-bp minimal ftz enhancer (see Figure S2 for sequence). Bent arrow indicates the start of the lacZ sequence. Vertical bars above and below the line represent positions of recognition sequences for restriction enzymes BstUI (CGCG) and HaeIII (GGCC), respectively. (B–E) Time courses of restriction digests of genomic DNA with enzymes sensitive (BstUI) or insensitive (HaeIII) to methylated DNA are shown as agarose gel pictures and quantifications. Genomic DNA was isolated from Kr 1 /CyO ftz-lacZ and +/CyO ftz-lacZ (wild-type control) adult flies (B, C), or from the F1 progeny flies of Kr 1 /CyO ftz-lacZ males crossed to hop Tum-l/+ females or wild-type females (D, E), and digested with the indicated enzymes for the indicated times (minutes). Digested DNA was amplified with PCR primers shown in (A) and run on an agarose gel. Note that the genomic DNA from Kr +/ − flies or from the F1 progeny of hop Tum-l/+ females and Kr 1 /CyO ftz-lacZ males is more resistant to BstUI digestion than the controls. (F, G) Digested genomic DNA purified from embryos was immunoprecipitated by antibodies to methylated cytosine and amplified by PCR primers shown in (A). Embryos derived from wild-type or Kr +/ − parents (F), or from Kr 1 /ftz-lacZ males crossed to wild-type (top) or Tum-l/+ (bottom) females (G) were used for DNA isolation. Note the presence of higher levels of 5-meC in the ftz-lacZ promoter in Kr +/ − embryos or in those from Tum-l/+ females crossed to Kr 1 /ftz-lacZ males.

    Techniques Used: Methylation, Expressing, Polymerase Chain Reaction, Sequencing, Agarose Gel Electrophoresis, Isolation, Amplification, Purification, Immunoprecipitation, Derivative Assay, DNA Extraction

    4) Product Images from "Selective amplification of hypermethylated DNA from diverse tumor types via MSRE-PCR"

    Article Title: Selective amplification of hypermethylated DNA from diverse tumor types via MSRE-PCR

    Journal: Oncotarget

    doi: 10.18632/oncotarget.27825

    Methylation sensitive restriction sites in target cDMRs. Five MSREs are considered here: HhaI, HpaII, HpyCH4IV, AciI and BstUI. ( A ) Histogram of the number of MSRE sites in the target regions. ( B ) Scatterplot of the number of MSRE sites versus the number of CpGs in the target regions showing a linear relationship (R 2 = 0.89, p
    Figure Legend Snippet: Methylation sensitive restriction sites in target cDMRs. Five MSREs are considered here: HhaI, HpaII, HpyCH4IV, AciI and BstUI. ( A ) Histogram of the number of MSRE sites in the target regions. ( B ) Scatterplot of the number of MSRE sites versus the number of CpGs in the target regions showing a linear relationship (R 2 = 0.89, p

    Techniques Used: Methylation

    5) Product Images from "In Vitro Grown Sheep Preantral Follicles Yield Oocytes with Normal Nuclear-Epigenetic Maturation"

    Article Title: In Vitro Grown Sheep Preantral Follicles Yield Oocytes with Normal Nuclear-Epigenetic Maturation

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0027550

    Methylation status of the IGF2R and BEGAIN genes in germ cells analyzed by bisulfite sequencing. An example of bisulphite sequencing analysis of examined CpGs in three imprinted genes. Each line represents an individual sequence molecule, with each circle corresponding to a separate CpG site (CpG). Black and white circles indicate methylated and unmethylated CpGs respectively. Arrows indicate the CpGs assayed by BstUI and HhaI enzymes, used in the combined bisulfite restriction analysis (COBRA). GV, germinal vesicle; MII, metaphase II; IVM, in vitro maturation; PA, preantral follicles; EA, early antral follicles; Preov, preovulatory follicles; A, antral follicles. Percentages of methylation were calculated by counting the number of methylated CpGs out of the total number of CpGs within the fragment amplified.
    Figure Legend Snippet: Methylation status of the IGF2R and BEGAIN genes in germ cells analyzed by bisulfite sequencing. An example of bisulphite sequencing analysis of examined CpGs in three imprinted genes. Each line represents an individual sequence molecule, with each circle corresponding to a separate CpG site (CpG). Black and white circles indicate methylated and unmethylated CpGs respectively. Arrows indicate the CpGs assayed by BstUI and HhaI enzymes, used in the combined bisulfite restriction analysis (COBRA). GV, germinal vesicle; MII, metaphase II; IVM, in vitro maturation; PA, preantral follicles; EA, early antral follicles; Preov, preovulatory follicles; A, antral follicles. Percentages of methylation were calculated by counting the number of methylated CpGs out of the total number of CpGs within the fragment amplified.

    Techniques Used: Methylation, Methylation Sequencing, Bisulfite Sequencing, Sequencing, Combined Bisulfite Restriction Analysis Assay, In Vitro, Amplification

    6) Product Images from "Population-Based Molecular Epidemiology of Leprosy in Cebu, Philippines "

    Article Title: Population-Based Molecular Epidemiology of Leprosy in Cebu, Philippines

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.02021-08

    The 21-3 and (GGT)5 alleles are indicative of SNP types 1 and 3 in Philippine M. leprae strains. The table indicates the VNTR alleles for 21-3 and (GGT)5 for 10 M. leprae specimens. The gel shows the corresponding SNP locus 3 BstUI cutting patterns for
    Figure Legend Snippet: The 21-3 and (GGT)5 alleles are indicative of SNP types 1 and 3 in Philippine M. leprae strains. The table indicates the VNTR alleles for 21-3 and (GGT)5 for 10 M. leprae specimens. The gel shows the corresponding SNP locus 3 BstUI cutting patterns for

    Techniques Used:

    7) Product Images from "Bisulfite-independent analysis of CpG island methylation enables genome-scale stratification of single cells"

    Article Title: Bisulfite-independent analysis of CpG island methylation enables genome-scale stratification of single cells

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkx026

    Schematic representation of CGI-seq. TEST: single cell being studied. Methylation control (MC): cells that are used to define the detectable CGIs or any other accessible methylated regions (AMRs), sharing the same genome as the TEST. For CGI-seq analysis, the genomic DNA is classified into three classes of sequences: Me-CGIs (representing methylated CGIs, and more generally, Me-AMRs) indicated as a red line, Um-CGIs (unmethylated CGIs, and generally Um-AMRs) indicated as a blue line and undetectable regions indicated as a black line. Green lines are amplified sequences. Solid dot: Me-CpG sites. Hollow dot: Um-CpG sites. The intact gDNA is released from cell (step 1) and digested with RE1 (TEST) or no digestion (MC) (step 2), followed by MDA amplification (step 3). The amplification product is then digested by RE2 (step 4), in which the short fragments are converted t‘o a library and subjected to massive sequencing (step 5). The reads are aligned to the genome (step 6) and the methylation status of the AMRs (particularly CGIs) is determined (‘Materials and Methods’ section). The AMRs detected in both the TEST and the MC are called Me-CGIs (or Me-AMRs). The AMRs uniquely detected in the MC but not in the TEST are called Um-CGIs (or Um-AMRs) of the TEST. RE1 (four MREs in combination: Cfr42I (SacII), PdiI (NaeI), Eco52I (EagI) and PteI(BssHII), FastDigest enzyme from Thermal Scientific Inc.) distinguishes Me-CGIs (Me-AMRs) from Um-CGIs (Um-AMRs). MDA selectively depletes Um-CGIs (Um-AMRs) of the TEST and amplifies Me-CGIs (Me-AMRs). RE2 (either BstUI alone, or a combination of 3 separate REs (ABH: AciI, BstUI and Hinp1I from NEB)) generates fragments specifically enriching CGI sequences (and other CG-rich regions) from non-CGI sequences.
    Figure Legend Snippet: Schematic representation of CGI-seq. TEST: single cell being studied. Methylation control (MC): cells that are used to define the detectable CGIs or any other accessible methylated regions (AMRs), sharing the same genome as the TEST. For CGI-seq analysis, the genomic DNA is classified into three classes of sequences: Me-CGIs (representing methylated CGIs, and more generally, Me-AMRs) indicated as a red line, Um-CGIs (unmethylated CGIs, and generally Um-AMRs) indicated as a blue line and undetectable regions indicated as a black line. Green lines are amplified sequences. Solid dot: Me-CpG sites. Hollow dot: Um-CpG sites. The intact gDNA is released from cell (step 1) and digested with RE1 (TEST) or no digestion (MC) (step 2), followed by MDA amplification (step 3). The amplification product is then digested by RE2 (step 4), in which the short fragments are converted t‘o a library and subjected to massive sequencing (step 5). The reads are aligned to the genome (step 6) and the methylation status of the AMRs (particularly CGIs) is determined (‘Materials and Methods’ section). The AMRs detected in both the TEST and the MC are called Me-CGIs (or Me-AMRs). The AMRs uniquely detected in the MC but not in the TEST are called Um-CGIs (or Um-AMRs) of the TEST. RE1 (four MREs in combination: Cfr42I (SacII), PdiI (NaeI), Eco52I (EagI) and PteI(BssHII), FastDigest enzyme from Thermal Scientific Inc.) distinguishes Me-CGIs (Me-AMRs) from Um-CGIs (Um-AMRs). MDA selectively depletes Um-CGIs (Um-AMRs) of the TEST and amplifies Me-CGIs (Me-AMRs). RE2 (either BstUI alone, or a combination of 3 separate REs (ABH: AciI, BstUI and Hinp1I from NEB)) generates fragments specifically enriching CGI sequences (and other CG-rich regions) from non-CGI sequences.

    Techniques Used: Methylation, Amplification, Multiple Displacement Amplification, Sequencing

    Characterization of CGI-seq on low quantity cells (LQC). ( A ) The correlation between the averaged methylation score of five 100-cell K562 samples and bulk M450K data with 350 000 K562 cells (UCSC ENCODE website). The Pearson correlation based on all informative CGIs was used to measure the diversity of CpG methylation. Region SD is the standard deviation of methylation scores of all CpGs in one CGI. Points in dark blue clusters are those CGIs whose CpGs have a similar methylation score, and points in light blue are those CGIs with diverse methylation of their CpGs. ( B ) DNA methylation levels of 41 classes of different regulatory markers between bulk ENCODE RRBS data and averaged LQC data generated by CGI-seq for K562 cell line. These 41 regions include Cbp, Cbx2, Cbx3, Cbx8, CGI, Chd1, Chd4, Chd7, Cfcf, Ezh2, H2az, H3k27ac, H3k27me3, H3k36me3, H3k4me1, H3k4me2, H3k4me3, H3k79me2, H3k9ac, H3k9me1, H3k9me3, H4k20me1, Hdac1, Hdac2, Hdac6, Lsd1, Ncor, Nsd2, P300, Pcaf, Phf8, Plu1, Pol2b, Promoter, Rbbp5, Rest, Rnf2, Sap30, Setdp1, Sirt6 and Suz12. ( C ) CGI-seq clustering based on Pearson correlation of CGI methylation across 30 LQC samples of four cell lines (Fibroblast-100C, GM12878-100C, iPS-100C, K562-100C and K562-10C). 100C: 100-cell sample; 10C: 10-cell sample. The samples not indicated with ABH (AciI, BstUI and HpaII) or BstUI are all cut by ABH as RE2. ( D ) Heatmap of the top 1000 variant CGIs in methylation across 30 LQC samples of four cell lines.
    Figure Legend Snippet: Characterization of CGI-seq on low quantity cells (LQC). ( A ) The correlation between the averaged methylation score of five 100-cell K562 samples and bulk M450K data with 350 000 K562 cells (UCSC ENCODE website). The Pearson correlation based on all informative CGIs was used to measure the diversity of CpG methylation. Region SD is the standard deviation of methylation scores of all CpGs in one CGI. Points in dark blue clusters are those CGIs whose CpGs have a similar methylation score, and points in light blue are those CGIs with diverse methylation of their CpGs. ( B ) DNA methylation levels of 41 classes of different regulatory markers between bulk ENCODE RRBS data and averaged LQC data generated by CGI-seq for K562 cell line. These 41 regions include Cbp, Cbx2, Cbx3, Cbx8, CGI, Chd1, Chd4, Chd7, Cfcf, Ezh2, H2az, H3k27ac, H3k27me3, H3k36me3, H3k4me1, H3k4me2, H3k4me3, H3k79me2, H3k9ac, H3k9me1, H3k9me3, H4k20me1, Hdac1, Hdac2, Hdac6, Lsd1, Ncor, Nsd2, P300, Pcaf, Phf8, Plu1, Pol2b, Promoter, Rbbp5, Rest, Rnf2, Sap30, Setdp1, Sirt6 and Suz12. ( C ) CGI-seq clustering based on Pearson correlation of CGI methylation across 30 LQC samples of four cell lines (Fibroblast-100C, GM12878-100C, iPS-100C, K562-100C and K562-10C). 100C: 100-cell sample; 10C: 10-cell sample. The samples not indicated with ABH (AciI, BstUI and HpaII) or BstUI are all cut by ABH as RE2. ( D ) Heatmap of the top 1000 variant CGIs in methylation across 30 LQC samples of four cell lines.

    Techniques Used: Methylation, CpG Methylation Assay, Standard Deviation, DNA Methylation Assay, Generated, Variant Assay

    8) Product Images from "Tissue-specific alternative polyadenylation at the imprinted gene Mest regulates allelic usage at Copg2"

    Article Title: Tissue-specific alternative polyadenylation at the imprinted gene Mest regulates allelic usage at Copg2

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkr871

    Allelic usage at Copg2 is regulated by MestXL , which is not nuclear localized. ( A ) Hot-stop PCR/RFLP analysis on various E14.5 F1 Mest C/+ ( 1 ) and Mest C/KO ( 2 ) tissues reveals allelic parental differences for Copg2 in tissues that express MestXL . 32 P-labeled Copg2 RT–PCR products were digested with BstUI, which cuts only the maternal CAST (C) allele, and ran out on an 8% polyacrylamide gel. Controls on paternal (CD-1) and maternal (C) RNA show complete digestion of the maternal CAST allele with BstUI. P:paternal allele; M:maternal allele. ( B ) Maternal and paternal band densities were calculated using Image J analysis software and an M/P ratio was generated for each sample. ( C ) Direct sequencing of three Copg2 SNPs for RNA isolated from four different tissues from 4 Mest C/+ and 6 Mest C/KO E14.5 embryos expressed as maternal to paternal (M/P) allele ratios. The phred program was used to read the sequence trace data and assign quality values to the bases ( 24 ) and the numbers were generated after normalizing to F1 genomic DNA. Differences between Mest C/+ and Mest C/KO for each E14.5 tissue were evaluated using the two-tailed unpaired Student t -test ( P -values) and those differences that were statistically significant are marked with an asterisk ( P
    Figure Legend Snippet: Allelic usage at Copg2 is regulated by MestXL , which is not nuclear localized. ( A ) Hot-stop PCR/RFLP analysis on various E14.5 F1 Mest C/+ ( 1 ) and Mest C/KO ( 2 ) tissues reveals allelic parental differences for Copg2 in tissues that express MestXL . 32 P-labeled Copg2 RT–PCR products were digested with BstUI, which cuts only the maternal CAST (C) allele, and ran out on an 8% polyacrylamide gel. Controls on paternal (CD-1) and maternal (C) RNA show complete digestion of the maternal CAST allele with BstUI. P:paternal allele; M:maternal allele. ( B ) Maternal and paternal band densities were calculated using Image J analysis software and an M/P ratio was generated for each sample. ( C ) Direct sequencing of three Copg2 SNPs for RNA isolated from four different tissues from 4 Mest C/+ and 6 Mest C/KO E14.5 embryos expressed as maternal to paternal (M/P) allele ratios. The phred program was used to read the sequence trace data and assign quality values to the bases ( 24 ) and the numbers were generated after normalizing to F1 genomic DNA. Differences between Mest C/+ and Mest C/KO for each E14.5 tissue were evaluated using the two-tailed unpaired Student t -test ( P -values) and those differences that were statistically significant are marked with an asterisk ( P

    Techniques Used: Polymerase Chain Reaction, Labeling, Reverse Transcription Polymerase Chain Reaction, Software, Generated, Sequencing, Isolation, Two Tailed Test

    9) Product Images from "Epigenetic Regulation of Dpp6 Expression by Dnmt3b and Its Novel Role in the Inhibition of RA Induced Neuronal Differentiation of P19 Cells"

    Article Title: Epigenetic Regulation of Dpp6 Expression by Dnmt3b and Its Novel Role in the Inhibition of RA Induced Neuronal Differentiation of P19 Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0055826

    Association of Dnmt3b with the promoter of Dpp6 gene, methylation pattern of Dpp6 promoter CpG Island and its expression in P19 cells. A, Recruitment of Dnmt1, Dnmt3a, and Dnmt3b on Dpp6 promoter in undifferentiated (P19 -RA) and differentiated P19 cells by RA treatment for initial 2 days and further culture for 4 days without RA by using quantitative ChIP analysis. Data was normalized to input fraction. The value of IgG (negative control) was set as 1, and the results were presented as relative fold enrichment of IgG. Values represent mean of three independent experiments and error bars represent ± SEM. ns = no significant difference between P19–RA and P19+RA. B, The sequence of 222 bp PCR product used for BGS, and COBRA. Arrows indicate the position of primers and 17 CpG dinucleotides included in this analysis are shown in bold. Restriction Sites for TaqI and BstUI enzymes are underlined. C, Bisulfite conversion control PCR by using primer sets that can distinguish between converted (Control 1) and unconverted DNA (Control 2). D, Bisulfite genomic sequence analysis of Dpp6 promoter in undifferentiated (P19 -RA) and differentiated P19 cells by RA treatment for initial 2 days and further culture for 4 days without RA. 17 CpG sites were analyzed in 15 individual clones. The red and blue colored boxes showed methylated and unmethylated cytosines respectively. E, For COBRA analysis, the PCR products were digested with TaqI and BstUI followed by agarose gel electrophoresis to analyze methylation status at Dpp6 promoter. F, Methylation Specific PCR was performed using bisulfite treated DNA and primers that can discriminate between unmethylated (U) and methylated (M) DNA. G, Quantitative real time RT-PCR analysis of Dpp6 in untreated P19 cells (D0) and RA treated P19 cells at different days (D2, D4, D6). Values represent mean of three independent experiments and error bars represent ± SEM. H, Western blot analysis showing protein expression of Dpp6 in RA untreated and RA treated P19 cells at different days.
    Figure Legend Snippet: Association of Dnmt3b with the promoter of Dpp6 gene, methylation pattern of Dpp6 promoter CpG Island and its expression in P19 cells. A, Recruitment of Dnmt1, Dnmt3a, and Dnmt3b on Dpp6 promoter in undifferentiated (P19 -RA) and differentiated P19 cells by RA treatment for initial 2 days and further culture for 4 days without RA by using quantitative ChIP analysis. Data was normalized to input fraction. The value of IgG (negative control) was set as 1, and the results were presented as relative fold enrichment of IgG. Values represent mean of three independent experiments and error bars represent ± SEM. ns = no significant difference between P19–RA and P19+RA. B, The sequence of 222 bp PCR product used for BGS, and COBRA. Arrows indicate the position of primers and 17 CpG dinucleotides included in this analysis are shown in bold. Restriction Sites for TaqI and BstUI enzymes are underlined. C, Bisulfite conversion control PCR by using primer sets that can distinguish between converted (Control 1) and unconverted DNA (Control 2). D, Bisulfite genomic sequence analysis of Dpp6 promoter in undifferentiated (P19 -RA) and differentiated P19 cells by RA treatment for initial 2 days and further culture for 4 days without RA. 17 CpG sites were analyzed in 15 individual clones. The red and blue colored boxes showed methylated and unmethylated cytosines respectively. E, For COBRA analysis, the PCR products were digested with TaqI and BstUI followed by agarose gel electrophoresis to analyze methylation status at Dpp6 promoter. F, Methylation Specific PCR was performed using bisulfite treated DNA and primers that can discriminate between unmethylated (U) and methylated (M) DNA. G, Quantitative real time RT-PCR analysis of Dpp6 in untreated P19 cells (D0) and RA treated P19 cells at different days (D2, D4, D6). Values represent mean of three independent experiments and error bars represent ± SEM. H, Western blot analysis showing protein expression of Dpp6 in RA untreated and RA treated P19 cells at different days.

    Techniques Used: Methylation, Expressing, Chromatin Immunoprecipitation, Negative Control, Sequencing, Polymerase Chain Reaction, Combined Bisulfite Restriction Analysis Assay, Clone Assay, Agarose Gel Electrophoresis, Quantitative RT-PCR, Western Blot

    10) Product Images from "Discovery of DNA methylation markers in cervical cancer using relaxation ranking"

    Article Title: Discovery of DNA methylation markers in cervical cancer using relaxation ranking

    Journal: BMC Medical Genomics

    doi: 10.1186/1755-8794-1-57

    Representative COBRA on 3 gene promoters ( SST , AUTS2 and SYCP3 ) . A: schematic representation of of the restriction enzyme sites in the virtual hypermethylated BSP nucleotide sequence after bisulfite treatment.(B: BstUI , T: TaqI and H: HinfI ). Bars represent CG site and arrow is TSS (retrieved from Ensembl). B: Result of COBRA analysis of BSP products of tumour samples (T1-T10) and 5 normal cervices (N1-N5), in vitro methylated DNA as a positive control (IV) and leukocyte DNA as a negative (unmethylated) control (L); lane B is water blank.
    Figure Legend Snippet: Representative COBRA on 3 gene promoters ( SST , AUTS2 and SYCP3 ) . A: schematic representation of of the restriction enzyme sites in the virtual hypermethylated BSP nucleotide sequence after bisulfite treatment.(B: BstUI , T: TaqI and H: HinfI ). Bars represent CG site and arrow is TSS (retrieved from Ensembl). B: Result of COBRA analysis of BSP products of tumour samples (T1-T10) and 5 normal cervices (N1-N5), in vitro methylated DNA as a positive control (IV) and leukocyte DNA as a negative (unmethylated) control (L); lane B is water blank.

    Techniques Used: Combined Bisulfite Restriction Analysis Assay, Sequencing, In Vitro, Methylation, Positive Control

    11) Product Images from "Detection of Cryptosporidium and Identification to the Species Level by Nested PCR and Restriction Fragment Length Polymorphism"

    Article Title: Detection of Cryptosporidium and Identification to the Species Level by Nested PCR and Restriction Fragment Length Polymorphism

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.43.3.1017-1023.2005

    Comparative restriction patterns of C. hominis (A), C. parvum (B), C. felis (C), and C. meleagridis (D) obtained with the following enzymes: TaqI (lanes 1), AseI (lanes 2), MseI (lanes 3), BstUI (lanes 4), and SspI (lanes 5). Lanes 6, undigested C. hominis
    Figure Legend Snippet: Comparative restriction patterns of C. hominis (A), C. parvum (B), C. felis (C), and C. meleagridis (D) obtained with the following enzymes: TaqI (lanes 1), AseI (lanes 2), MseI (lanes 3), BstUI (lanes 4), and SspI (lanes 5). Lanes 6, undigested C. hominis

    Techniques Used:

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    Article Snippet: Wild and bisulfite-modified DNAs were amplified by an initial incubation at 94°C for 2 min followed by 30 cycles (for wild DNA) or 40 cycles (for modified DNA) of 94°C for 30 s, 65°C for 30 s, 72°C for 1 min, and a final extension at 72°C for 10 min using Platinum PCR SuperMix (Invitrogen). .. Aliquots (30 μL) of the PCR-amplified products were digested with restriction enzymes, 40 units of BstU I at 60°C for 4 h (5′…cg^cg…3′; New England Biolabs, Beverly, MA, USA). .. Then, the products were subjected to electrophoresis on a 3% agarose gel and visualized by SYBR Green I (BMA).

    Article Title: Homozygosity for Pro of p53 Arg72Pro as a potential risk factor for hepatocellular carcinoma in Chinese population
    Article Snippet: The PCR primers used for amplifying the polymorphism region were: forward, 5’-TTGCCGTCCCAAGCAATGGATGA-3’; reverse, 5’-TCTGGGAAGGGACAGAAGATGAC-3’. .. PCR condition was 2 min at 94 °C, followed by 35 cycles of 30 s at 94 °C, 30 s at 60 °C, and 30 s at 72 °C, and with a final extension at 72 °C for 7 min. A 10-μL aliquot of PCR product was digested overnight at 60 °C in a 15-μL reaction volume containing 10 units of BstU I (New England BioLabs). .. After overnight digestion, the fragments were separated by electrophoresis on a vertical 90 g/L non-denaturing polyacrylamide gel at 120 V for 45 min, stained with ethidium bromide.

    Article Title: Recurrent mutations in a SERPINC1 hotspot associate with venous thrombosis without apparent antithrombin deficiency
    Article Snippet: We therefore amplified exon 5 of SERPINC1 using the following oligonucleotide primers: forward primer, 5’-TCATTCTGACACAGCCATT-3’; and reverse primer, 5’-CCTGACTTGTTGCTCCTTT-3’. .. Identification of the c.883G > A mutation was done by restriction analysis of the 649-bp PCR product with BstUI (New England Biolabs, Ipswich, MA, USA). .. Briefly, 10 μL PCR product was completely digested with 2 U of BstUI during 2 hours at 60°C.

    Article Title: Lack of HLA class II antigen expression in microsatellite unstable colorectal carcinomas is caused by mutations in HLA class II regulatory genes
    Article Snippet: Primers for methylation analysis and PCR conditions were used as described . .. The PCR products were digested with BstUI (New England Biolabs, Frankfurt, Germany). .. DNA from the cell line HCT116 was included as a positive control for CIITA promoter IV methylation in each assay.

    Incubation:

    Article Title: Evidence for Transgenerational Transmission of Epigenetic Tumor Susceptibility in Drosophila
    Article Snippet: The samples were treated with DNase-free RNase A (Sigma) for 2 h at 37 °C prior to column purification. .. For restriction digests, 3 μg of genomic DNA was incubated with 10 units of BstUI (New England Biolabs, http://www.neb.com/ ) or 10 units of HaeIII (New England Biolabs) in 60 μl of total volume at 37 °C. ..

    Mutagenesis:

    Article Title: Recurrent mutations in a SERPINC1 hotspot associate with venous thrombosis without apparent antithrombin deficiency
    Article Snippet: We therefore amplified exon 5 of SERPINC1 using the following oligonucleotide primers: forward primer, 5’-TCATTCTGACACAGCCATT-3’; and reverse primer, 5’-CCTGACTTGTTGCTCCTTT-3’. .. Identification of the c.883G > A mutation was done by restriction analysis of the 649-bp PCR product with BstUI (New England Biolabs, Ipswich, MA, USA). .. Briefly, 10 μL PCR product was completely digested with 2 U of BstUI during 2 hours at 60°C.

    Agarose Gel Electrophoresis:

    Article Title: Human Topoisomerase I Promotes Initiation of Simian Virus 40 DNA Replication In Vitro
    Article Snippet: Topo I-deficient 293 cell extracts were incubated with T antigen and pSV011+ in replication buffer B (40 mM HEPES-KOH [pH 7.5], 8 mM MgCl2 , 0.5 mM DTT, 3 mM ATP, 40 mM creatine phosphate, 20 to 40 ng of creatine phosphokinase per μl [final concentrations]) at 37°C for 30 min. To initiate DNA synthesis, GTP, CTP, and UTP (0.2 mM each), dGTP, dCTP, and dTTP (0.1 mM each), and 1 μCi of [α-32 P]dATP (3,000 Ci/mmol) were added with or without topo I, and DNA synthesis continued at 37°C for 30 min. .. In some cases, after separation of the replication products on a 1.5% agarose gel, the completed molecules were visualized by ethidium bromide, excised from the gel, Cerenkov counted, purified by GeneClean (Bio 101), digested with Bst UI and Ssp I (New England Biolabs), and applied to a 2% agarose gel. .. Pulse-chase assays were performed by a modification of the procedures described by Fotedar et al. ( ) (see Fig. A).

    Purification:

    Article Title: Human Topoisomerase I Promotes Initiation of Simian Virus 40 DNA Replication In Vitro
    Article Snippet: Topo I-deficient 293 cell extracts were incubated with T antigen and pSV011+ in replication buffer B (40 mM HEPES-KOH [pH 7.5], 8 mM MgCl2 , 0.5 mM DTT, 3 mM ATP, 40 mM creatine phosphate, 20 to 40 ng of creatine phosphokinase per μl [final concentrations]) at 37°C for 30 min. To initiate DNA synthesis, GTP, CTP, and UTP (0.2 mM each), dGTP, dCTP, and dTTP (0.1 mM each), and 1 μCi of [α-32 P]dATP (3,000 Ci/mmol) were added with or without topo I, and DNA synthesis continued at 37°C for 30 min. .. In some cases, after separation of the replication products on a 1.5% agarose gel, the completed molecules were visualized by ethidium bromide, excised from the gel, Cerenkov counted, purified by GeneClean (Bio 101), digested with Bst UI and Ssp I (New England Biolabs), and applied to a 2% agarose gel. .. Pulse-chase assays were performed by a modification of the procedures described by Fotedar et al. ( ) (see Fig. A).

    Generated:

    Article Title: High-Risk Human Papillomavirus E7 Alters Host DNA Methylome and Represses HLA-E Expression in Human Keratinocytes
    Article Snippet: MSP was performed with primers described in Table and validated using methylated or unmethylated control gDNA. .. Control DNA was generated by in vitro DNA methylation of gDNA extracted from W12E, W12G, and W12GPXY cells using McrBC endonuclease or M.SssI CpG methyltransferase followed by BstUI digestion (New England Biolabs). .. In vitro DNA methylation was performed using the M.SssI CpG methyltransferase and methylation efficiency was validated by McrBC or BstUI digestion.

    In Vitro:

    Article Title: High-Risk Human Papillomavirus E7 Alters Host DNA Methylome and Represses HLA-E Expression in Human Keratinocytes
    Article Snippet: MSP was performed with primers described in Table and validated using methylated or unmethylated control gDNA. .. Control DNA was generated by in vitro DNA methylation of gDNA extracted from W12E, W12G, and W12GPXY cells using McrBC endonuclease or M.SssI CpG methyltransferase followed by BstUI digestion (New England Biolabs). .. In vitro DNA methylation was performed using the M.SssI CpG methyltransferase and methylation efficiency was validated by McrBC or BstUI digestion.

    DNA Methylation Assay:

    Article Title: High-Risk Human Papillomavirus E7 Alters Host DNA Methylome and Represses HLA-E Expression in Human Keratinocytes
    Article Snippet: MSP was performed with primers described in Table and validated using methylated or unmethylated control gDNA. .. Control DNA was generated by in vitro DNA methylation of gDNA extracted from W12E, W12G, and W12GPXY cells using McrBC endonuclease or M.SssI CpG methyltransferase followed by BstUI digestion (New England Biolabs). .. In vitro DNA methylation was performed using the M.SssI CpG methyltransferase and methylation efficiency was validated by McrBC or BstUI digestion.

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    New England Biolabs bstui
    Analysis of CIITA promoter IV methylation in MSI-H CRC lesions (A) and cell lines (B). Two regions (GM1 and GM2) of the promoter were analyzed for each sample. Methylation was analyzed by enzymatic digest with <t>BstUI</t> following bisulfite treatment and <t>PCR</t>
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    Analysis of CIITA promoter IV methylation in MSI-H CRC lesions (A) and cell lines (B). Two regions (GM1 and GM2) of the promoter were analyzed for each sample. Methylation was analyzed by enzymatic digest with BstUI following bisulfite treatment and PCR

    Journal: International journal of cancer. Journal international du cancer

    Article Title: Lack of HLA class II antigen expression in microsatellite unstable colorectal carcinomas is caused by mutations in HLA class II regulatory genes

    doi: 10.1002/ijc.25106

    Figure Lengend Snippet: Analysis of CIITA promoter IV methylation in MSI-H CRC lesions (A) and cell lines (B). Two regions (GM1 and GM2) of the promoter were analyzed for each sample. Methylation was analyzed by enzymatic digest with BstUI following bisulfite treatment and PCR

    Article Snippet: The PCR products were digested with BstUI (New England Biolabs, Frankfurt, Germany).

    Techniques: Methylation, Polymerase Chain Reaction

    Identification of a mutational hotspot in SERPINC1 (A) Electropherogram of samples carrying all mutations affecting antithrombin Arg294 and Val295 residues. (B) Restriction fragments generated with BstUI on the PCR product of SERPINC1 exon 5 using the primers described in Materials and Methods. (C) Localization in the structure of antithrombin of affected residues in the native, activated, and latent configurations. The central A sheet is colored in blue, the reactive center loop in red, and s1B in green. Model building was performed by using SWISS-MODEL and the Swiss-PdbViewer programs [Guex N, Peitsch MC. SWISS-MODEL and the Swiss-PdbViewer: an environment for comparative protein modelling. Electrophoresis 1997; 18: 2714–23] ( http://www.expasy.ch/spdbv ).

    Journal: Oncotarget

    Article Title: Recurrent mutations in a SERPINC1 hotspot associate with venous thrombosis without apparent antithrombin deficiency

    doi: 10.18632/oncotarget.21365

    Figure Lengend Snippet: Identification of a mutational hotspot in SERPINC1 (A) Electropherogram of samples carrying all mutations affecting antithrombin Arg294 and Val295 residues. (B) Restriction fragments generated with BstUI on the PCR product of SERPINC1 exon 5 using the primers described in Materials and Methods. (C) Localization in the structure of antithrombin of affected residues in the native, activated, and latent configurations. The central A sheet is colored in blue, the reactive center loop in red, and s1B in green. Model building was performed by using SWISS-MODEL and the Swiss-PdbViewer programs [Guex N, Peitsch MC. SWISS-MODEL and the Swiss-PdbViewer: an environment for comparative protein modelling. Electrophoresis 1997; 18: 2714–23] ( http://www.expasy.ch/spdbv ).

    Article Snippet: Identification of the c.883G > A mutation was done by restriction analysis of the 649-bp PCR product with BstUI (New England Biolabs, Ipswich, MA, USA).

    Techniques: Generated, Polymerase Chain Reaction, Electrophoresis

    Non-denaturing polyacrylamide gel electrophoresis of p53 Arg72Pro region PCR products digested with BstU I . M: 100-bp DNA ladder, 1: Arg/Pro heterozygote, 2: Pro homozygote, 3: Arg homozygote.

    Journal:

    Article Title: Homozygosity for Pro of p53 Arg72Pro as a potential risk factor for hepatocellular carcinoma in Chinese population

    doi: 10.3748/wjg.v11.i2.289

    Figure Lengend Snippet: Non-denaturing polyacrylamide gel electrophoresis of p53 Arg72Pro region PCR products digested with BstU I . M: 100-bp DNA ladder, 1: Arg/Pro heterozygote, 2: Pro homozygote, 3: Arg homozygote.

    Article Snippet: PCR condition was 2 min at 94 °C, followed by 35 cycles of 30 s at 94 °C, 30 s at 60 °C, and 30 s at 72 °C, and with a final extension at 72 °C for 7 min. A 10-μL aliquot of PCR product was digested overnight at 60 °C in a 15-μL reaction volume containing 10 units of BstU I (New England BioLabs).

    Techniques: Polyacrylamide Gel Electrophoresis, Polymerase Chain Reaction

    Promoter CpG methylation correlates with lack of TUSC3 gene expression. (A) Schematic of TUSC3 locus showing the regions investigated by bisulfite pyrosequencing on the promoter (−105 to −57 relative to the transcriptional start site according to NM_006765) and genotyping assays of the 5′ untranslated region. PCR primers for DNA genotyping are indicated by black arrows while RT-PCR primers for mRNA genotyping are indicated by arrows highlighted in white. Enzyme recognition sites for Bst UI are indicated by “ B ”. (B) Methylation status of TUSC3 promoter region studied by bisulfite pyrosequencing. A similar methylation level of every CpG within each sample is observed and the gene follows “on-or-off” methylation pattern. Each circle represents a CpG site in a sample. Area shaded in black is proportional to the methylation level of the CpG site indicated by pyrosequencing. (C) Validation of complete methylation-sensitive restriction enzyme digestion on unmethylated molecules. Genomic DNA was predigested with Bst UI followed by PCR amplification with TUSC3 and ID2 specific primers ( Table S2 ). Bst UI digestion sites within the ID2 region were unmethylated ( Figure S6 ) and, therefore, no PCR product was generated after enzyme digestion. (D) Allele-specific methylation of TUSC3 on the fragment containing SNP rs12550009 demonstrated by enzyme digestion pyrosequencing. The “Simplex” diagrams (top) show the reference pyrograms by genotype. A heterozygous CT in the methylated samples (PM55 and PM123) displays a homozygous T pattern after Bst UI digestion indicating predominant methylation of the T allele. Allele-specific mRNA expression is concordant with allele-specific methylation on the same SNP rs1250009. Predominant expression of C alleles was observed in the cDNAs generated by RNA specific primers (bottom). RT+ and RT− represent assays with Reverse Transcriptase and without Reverse Transcriptase, respectively.

    Journal: PLoS ONE

    Article Title: Human Placental-Specific Epipolymorphism and its Association with Adverse Pregnancy Outcomes

    doi: 10.1371/journal.pone.0007389

    Figure Lengend Snippet: Promoter CpG methylation correlates with lack of TUSC3 gene expression. (A) Schematic of TUSC3 locus showing the regions investigated by bisulfite pyrosequencing on the promoter (−105 to −57 relative to the transcriptional start site according to NM_006765) and genotyping assays of the 5′ untranslated region. PCR primers for DNA genotyping are indicated by black arrows while RT-PCR primers for mRNA genotyping are indicated by arrows highlighted in white. Enzyme recognition sites for Bst UI are indicated by “ B ”. (B) Methylation status of TUSC3 promoter region studied by bisulfite pyrosequencing. A similar methylation level of every CpG within each sample is observed and the gene follows “on-or-off” methylation pattern. Each circle represents a CpG site in a sample. Area shaded in black is proportional to the methylation level of the CpG site indicated by pyrosequencing. (C) Validation of complete methylation-sensitive restriction enzyme digestion on unmethylated molecules. Genomic DNA was predigested with Bst UI followed by PCR amplification with TUSC3 and ID2 specific primers ( Table S2 ). Bst UI digestion sites within the ID2 region were unmethylated ( Figure S6 ) and, therefore, no PCR product was generated after enzyme digestion. (D) Allele-specific methylation of TUSC3 on the fragment containing SNP rs12550009 demonstrated by enzyme digestion pyrosequencing. The “Simplex” diagrams (top) show the reference pyrograms by genotype. A heterozygous CT in the methylated samples (PM55 and PM123) displays a homozygous T pattern after Bst UI digestion indicating predominant methylation of the T allele. Allele-specific mRNA expression is concordant with allele-specific methylation on the same SNP rs1250009. Predominant expression of C alleles was observed in the cDNAs generated by RNA specific primers (bottom). RT+ and RT− represent assays with Reverse Transcriptase and without Reverse Transcriptase, respectively.

    Article Snippet: For Bst UI predigestion assay followed by pyrosequencing on TUSC3 , 200 ng of genomic DNA was digested with 100 units of Bst UI (New England Biolabs) for 18 hours.

    Techniques: CpG Methylation Assay, Expressing, Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, Methylation, Amplification, Generated