hpa ii New England Biolabs Search Results


  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 95
    New England Biolabs hpa ii methyltransferase
    The Klf1 exon 2 CGI is unmethylated during development. A ) Schematic of the mouse Klf1 gene across exon 2 and the flanking introns shows salient restriction enzyme sites within the region and a 1.1-kbp cDNA probe used for Southern blot. B ) Genomic DNA samples from whole testis between P5 and P45 (lanes 1–7, left and center panels) and P45 spleen (lane 8). DNAs were purified and double-digested with Eco RI and Bam HI restriction enzymes to liberate the 1.85-kb genomic fragment containing the Klf1 exon 2 CGI. Thereafter, the DNA samples were further digested with <t>Hpa</t> II (left panel) or not digested (center panel) and then Southern blotted using the 1.1-kb probe shown in A . In addition, several samples were methylated using Hpa II <t>-methyltransferase</t> then digested with Hpa II (right panel) and Southern blotted. Because of the total number of samples in this experiment, three Southern blots were generated, and the relevant lanes were used to form composite panels. Data for the P5–P15 lanes (break in Southern blots) were obtained from separate Southern blots. Arrowheads in A , locations of Hpa II restriction sites; FL in B , full-length 1.85-kb Eco RI - Bam HI fragments that are visible in the center and right panels; Spl, spleen.
    Hpa Ii Methyltransferase, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 40 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hpa ii methyltransferase/product/New England Biolabs
    Average 95 stars, based on 40 article reviews
    Price from $9.99 to $1999.99
    hpa ii methyltransferase - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    95
    Millipore 5 aza 2
    The Klf1 exon 2 CGI is unmethylated during development. A ) Schematic of the mouse Klf1 gene across exon 2 and the flanking introns shows salient restriction enzyme sites within the region and a 1.1-kbp cDNA probe used for Southern blot. B ) Genomic DNA samples from whole testis between P5 and P45 (lanes 1–7, left and center panels) and P45 spleen (lane 8). DNAs were purified and double-digested with Eco RI and Bam HI restriction enzymes to liberate the 1.85-kb genomic fragment containing the Klf1 exon 2 CGI. Thereafter, the DNA samples were further digested with <t>Hpa</t> II (left panel) or not digested (center panel) and then Southern blotted using the 1.1-kb probe shown in A . In addition, several samples were methylated using Hpa II <t>-methyltransferase</t> then digested with Hpa II (right panel) and Southern blotted. Because of the total number of samples in this experiment, three Southern blots were generated, and the relevant lanes were used to form composite panels. Data for the P5–P15 lanes (break in Southern blots) were obtained from separate Southern blots. Arrowheads in A , locations of Hpa II restriction sites; FL in B , full-length 1.85-kb Eco RI - Bam HI fragments that are visible in the center and right panels; Spl, spleen.
    5 Aza 2, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 863 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/5 aza 2/product/Millipore
    Average 95 stars, based on 863 article reviews
    Price from $9.99 to $1999.99
    5 aza 2 - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    95
    New England Biolabs hpa ii
    Methylation analysis of the ZPBP gene in whole blood samples of Berkshire sows by PMP assay. gDNAs were cut with the methylation-sensitive restriction enzymes <t>Hpa</t> II/ <t>Msp</t> I. M: size marker; U: undigested DNA; SLG: smaller litter size group; LLG: larger litter size group.
    Hpa Ii, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 914 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hpa ii/product/New England Biolabs
    Average 95 stars, based on 914 article reviews
    Price from $9.99 to $1999.99
    hpa ii - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    95
    New England Biolabs hpa i
    <t>pRF/FMDV-IRES</t> and pCAGGS/FMDV-IRES plasmid construction. Structure of the bicistronic luciferase reporter construct containing the FMDV-IRES element located between Renilla luciferase and firefly luciferase (pRF-FMDV-IRS). Reporter gene was excised from this plasmid construct using the restriction enzymes Eco RV and Hpa I, and was o the pCAGGS/MCS(F) vector treated with Sma I and rAPid Alkaline Phosphatase using Mighty Mix
    Hpa I, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 188 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hpa i/product/New England Biolabs
    Average 95 stars, based on 188 article reviews
    Price from $9.99 to $1999.99
    hpa i - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    87
    New England Biolabs msp
    Examples of MSAP profiles showing various types of locus-specific DNA methylation inheritance and variation in F 1 hybrid and their parents (a) and different types of locus-specific DNA methylation inheritance and variation in F 4 and F 10 , and the F 4 and F 10 DNA methylation patterns (b,c) . (a). Primer combination is Eco RI+ ATT/ <t>Hpa</t> II ( <t>Msp</t> I) + ACC. Amplification results of Hpa II-digested genomic DNA of R. sativus L. (Lane 1), B. alboglabra Bailey (Lane 3) and F 1 hybrids (Lane 5) Amplification results of Msp I-digested genomic DNA of R. sativus L. (Lane 2), B. alboglabra Bailey (Lane 4), and F 1 hybrids (Lane 6). (b) and (c). Primer combinations are Eco RI + AGG/ Hpa II ( Msp I) + GAT and Eco RI + AGG/ Hpa II ( Msp I) + TCG, respectively. Arrows show variations of the various methylation patterns. H: Hpa II, M: Msp I.
    Msp, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 87/100, based on 97 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/msp/product/New England Biolabs
    Average 87 stars, based on 97 article reviews
    Price from $9.99 to $1999.99
    msp - by Bioz Stars, 2020-02
    87/100 stars
      Buy from Supplier

    95
    New England Biolabs endonuclease
    Examples of MSAP profiles showing various types of locus-specific DNA methylation inheritance and variation in F 1 hybrid and their parents (a) and different types of locus-specific DNA methylation inheritance and variation in F 4 and F 10 , and the F 4 and F 10 DNA methylation patterns (b,c) . (a). Primer combination is Eco RI+ ATT/ <t>Hpa</t> II ( <t>Msp</t> I) + ACC. Amplification results of Hpa II-digested genomic DNA of R. sativus L. (Lane 1), B. alboglabra Bailey (Lane 3) and F 1 hybrids (Lane 5) Amplification results of Msp I-digested genomic DNA of R. sativus L. (Lane 2), B. alboglabra Bailey (Lane 4), and F 1 hybrids (Lane 6). (b) and (c). Primer combinations are Eco RI + AGG/ Hpa II ( Msp I) + GAT and Eco RI + AGG/ Hpa II ( Msp I) + TCG, respectively. Arrows show variations of the various methylation patterns. H: Hpa II, M: Msp I.
    Endonuclease, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 30 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/endonuclease/product/New England Biolabs
    Average 95 stars, based on 30 article reviews
    Price from $9.99 to $1999.99
    endonuclease - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    87
    New England Biolabs hpa ii methylase
    Methylation states of the 5′ regions of the leukosialin gene exogenously introduced and stably maintained in non-leukosialin-expressing HeLa cells. (A) Schematic representations of the leukosialin gene and leukosialin <t>CAT</t> constructs. The exons are depicted by filled boxes, and introns and the 5′ flanking regions are depicted with horizontal lines. The vector sequences are depicted with dotted lines. Msp I (CCGG) sites are shown with vertical bars. The asterisk indicates the polymorphic site, where an Msp I site is lost in one allele of HeLa cells. The Msp I DNA fragment (560 bp) used for a hybridization probe is presented at the top. (B) Genomic DNAs (10 μg) from HeLa cells stably transfected with CpG-methylated- or unmethylated-leukosialin CAT constructs were digested with <t>Hpa</t> II (H) or Msp I (M) and separated by 1.5% agarose gel electrophoresis. The blotted filter was hybridized with the 560-bp DNA fragment of the 5′ region of the leukosialin gene shown in panel A. The hatched arrow indicates the position of the fragments produced by the endogenous gene as well as the exogenously introduced gene. The filled arrow indicates the signal produced by the exogenously introduced gene. The open arrow indicates the position of a polymorphic fragment of the endogenous leukosialin gene of HeLa cells.
    Hpa Ii Methylase, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 87/100, based on 51 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hpa ii methylase/product/New England Biolabs
    Average 87 stars, based on 51 article reviews
    Price from $9.99 to $1999.99
    hpa ii methylase - by Bioz Stars, 2020-02
    87/100 stars
      Buy from Supplier

    79
    New England Biolabs enzyme m hpa ii
    DNMT3L stimulates de novo methylation by Dnmt3a but not by DNMT3B at the SNRPN IC. <t>DNA</t> harvested from human 293/EBNA1 cells carrying pFC19 or pFC30 (in which a key ≈1-kb region of the SNRPN IC was cloned in either orientation) was digested with an excess of the methylation-sensitive enzyme <t>Hpa</t> II. The resulting DNA species were separated by electrophoresis through an agarose gel, transferred to a nylon membrane, and hybridized with a probe corresponding to the SNRPN IC. Two bands (684 and 754 bp for pFC19, and 684 and 221 bp for pFC30) were expected if no methylation was present. CpG methylation prevents cleavage by Hpa II and results in higher molecular weight species that can be easily separated. ( A ) Each target plasmid was introduced into cells alone, together with an expression vector for Dnmt3a, or together with expression vectors for both Dnmt3a and DNMT3L, as indicated. ( B ) Plasmid pFC19 was introduced in cells either with DNMT3B or with both DNMT3B and DNMT3L expression vectors, as indicated.
    Enzyme M Hpa Ii, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 79/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/enzyme m hpa ii/product/New England Biolabs
    Average 79 stars, based on 18 article reviews
    Price from $9.99 to $1999.99
    enzyme m hpa ii - by Bioz Stars, 2020-02
    79/100 stars
      Buy from Supplier

    82
    New England Biolabs restriction enzyme hpa ii
    ) were incubated the methylation-sensitive restriction enzyme <t>Hpa</t> II, which cleaves only unmethylated DNA; the PCR products then were subjected to capillary gel electrophoresis. The boxed numbers represent the area under each peak (ar), the peak height (ht), and the length (sz) of the DNA fragment. Note that peak area and height for the PCR products derived from individual <t>S2-II/6</t> are reduced in comparison to the amplicon derived from S2-II/5, indicating a partial loss of methylation.
    Restriction Enzyme Hpa Ii, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 82/100, based on 48 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/restriction enzyme hpa ii/product/New England Biolabs
    Average 82 stars, based on 48 article reviews
    Price from $9.99 to $1999.99
    restriction enzyme hpa ii - by Bioz Stars, 2020-02
    82/100 stars
      Buy from Supplier

    95
    New England Biolabs msp i
    Flowchart of MSD-library preparation. Genomic DNA (100 ng) was digested with 10 units of the primary restriction enzyme Sbf I for 1 h and then ligated with 0.5 nmol Adaptor A using 400 units of T4 DNA ligase for 2 h. The treated sample was then digested with 100 units of the non-methylation-sensitive restriction enzyme <t>Msp</t> I (100 units) followed by ligation of the ends of the DNA fragment with Adaptor B. The ligated DNA fragments were then digested with 50 units of Hpa II for 1 h. Owing to the methylation sensitivity of Hap II, only DNA fragments with a methylated <t>CpG</t> retained Adaptor B, which was removed from all other fragments. The DNA fragments were then subjected to Pre-PCR using specific primers for Adaptor A and Adaptor B. Fragments that did not contain Adaptor B at this stage were not amplified. The Pre-PCR amplicons (MSD library) were then amplified as a subpopulation by selective-PCR with 6-carboxyfluorescein (6-FAM)-labelled selective-PCR primers. Finally, the selective-PCR products were electrophoresed with a capillary sequencer and separated by length
    Msp I, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 1029 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/msp i/product/New England Biolabs
    Average 95 stars, based on 1029 article reviews
    Price from $9.99 to $1999.99
    msp i - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    78
    New England Biolabs cpg methylation sensitive restriction enzyme hpa ii
    Flowchart of MSD-library preparation. Genomic DNA (100 ng) was digested with 10 units of the primary restriction enzyme Sbf I for 1 h and then ligated with 0.5 nmol Adaptor A using 400 units of T4 DNA ligase for 2 h. The treated sample was then digested with 100 units of the non-methylation-sensitive restriction enzyme <t>Msp</t> I (100 units) followed by ligation of the ends of the DNA fragment with Adaptor B. The ligated DNA fragments were then digested with 50 units of Hpa II for 1 h. Owing to the methylation sensitivity of Hap II, only DNA fragments with a methylated <t>CpG</t> retained Adaptor B, which was removed from all other fragments. The DNA fragments were then subjected to Pre-PCR using specific primers for Adaptor A and Adaptor B. Fragments that did not contain Adaptor B at this stage were not amplified. The Pre-PCR amplicons (MSD library) were then amplified as a subpopulation by selective-PCR with 6-carboxyfluorescein (6-FAM)-labelled selective-PCR primers. Finally, the selective-PCR products were electrophoresed with a capillary sequencer and separated by length
    Cpg Methylation Sensitive Restriction Enzyme Hpa Ii, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 78/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cpg methylation sensitive restriction enzyme hpa ii/product/New England Biolabs
    Average 78 stars, based on 6 article reviews
    Price from $9.99 to $1999.99
    cpg methylation sensitive restriction enzyme hpa ii - by Bioz Stars, 2020-02
    78/100 stars
      Buy from Supplier

    78
    New England Biolabs restriction enzyme library hpa i
    Flowchart of MSD-library preparation. Genomic DNA (100 ng) was digested with 10 units of the primary restriction enzyme Sbf I for 1 h and then ligated with 0.5 nmol Adaptor A using 400 units of T4 DNA ligase for 2 h. The treated sample was then digested with 100 units of the non-methylation-sensitive restriction enzyme <t>Msp</t> I (100 units) followed by ligation of the ends of the DNA fragment with Adaptor B. The ligated DNA fragments were then digested with 50 units of Hpa II for 1 h. Owing to the methylation sensitivity of Hap II, only DNA fragments with a methylated <t>CpG</t> retained Adaptor B, which was removed from all other fragments. The DNA fragments were then subjected to Pre-PCR using specific primers for Adaptor A and Adaptor B. Fragments that did not contain Adaptor B at this stage were not amplified. The Pre-PCR amplicons (MSD library) were then amplified as a subpopulation by selective-PCR with 6-carboxyfluorescein (6-FAM)-labelled selective-PCR primers. Finally, the selective-PCR products were electrophoresed with a capillary sequencer and separated by length
    Restriction Enzyme Library Hpa I, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 78/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/restriction enzyme library hpa i/product/New England Biolabs
    Average 78 stars, based on 10 article reviews
    Price from $9.99 to $1999.99
    restriction enzyme library hpa i - by Bioz Stars, 2020-02
    78/100 stars
      Buy from Supplier

    81
    New England Biolabs hpa ii restriction endonuclease
    Resistance of methylated B. bovis DNA to cleavage by <t>Hpa</t> II. DNAs prepared from E. coli or B. bovis were treated with <t>Sss</t> I methylase and compared with untreated DNA for sensitivity to Hpa II digestion. DNA was visualized after electrophoresis on ethidium bromide-stained agarose gels. Lanes: M, 1-kb markers; 1, untreated DNA; 2, untreated DNA incubated with Hpa II; 3, methylated DNA; 4, methylated DNA incubated with Hpa II.
    Hpa Ii Restriction Endonuclease, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 81/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hpa ii restriction endonuclease/product/New England Biolabs
    Average 81 stars, based on 18 article reviews
    Price from $9.99 to $1999.99
    hpa ii restriction endonuclease - by Bioz Stars, 2020-02
    81/100 stars
      Buy from Supplier

    80
    New England Biolabs enzyme hpa i
    Genetic characterization Trypanosoma cruzi and Trypanosoma rangeli strains. (A) kDNA analysis of T . rangeli strains in a silver stained 6% polyacrylamide gel containing PCR products obtained with primers S35/S36/KP1L. The presence of a 165-bp band indicates the presence of KP1 minicircles. (B) Detection of the 832-bp fragment of TcSC5D gene in T . cruzi strains and clone. (C) PCR–RFLP of TcSC5D products digested with <t>Hpa</t> I and Sph I enzymes. MM: Molecular marker 100bp. NTC: No-template control.
    Enzyme Hpa I, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 80/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/enzyme hpa i/product/New England Biolabs
    Average 80 stars, based on 4 article reviews
    Price from $9.99 to $1999.99
    enzyme hpa i - by Bioz Stars, 2020-02
    80/100 stars
      Buy from Supplier

    79
    New England Biolabs isoschizomeric pair hpa ii
    Genetic characterization Trypanosoma cruzi and Trypanosoma rangeli strains. (A) kDNA analysis of T . rangeli strains in a silver stained 6% polyacrylamide gel containing PCR products obtained with primers S35/S36/KP1L. The presence of a 165-bp band indicates the presence of KP1 minicircles. (B) Detection of the 832-bp fragment of TcSC5D gene in T . cruzi strains and clone. (C) PCR–RFLP of TcSC5D products digested with <t>Hpa</t> I and Sph I enzymes. MM: Molecular marker 100bp. NTC: No-template control.
    Isoschizomeric Pair Hpa Ii, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 79/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/isoschizomeric pair hpa ii/product/New England Biolabs
    Average 79 stars, based on 8 article reviews
    Price from $9.99 to $1999.99
    isoschizomeric pair hpa ii - by Bioz Stars, 2020-02
    79/100 stars
      Buy from Supplier

    95
    New England Biolabs bst ui
    Methylation statuses at CpG sites near the RP2 onshore tandem GAAA repeat. Random ( A ) and non-random ( B ) X-inactivation patterns generated for different CpG-containing 5 me CpG-sensitive restriction endonuclease sites obtained using the 5 me CpG-based PCR RP2 / AR biplex assay across the restriction sites. Electropherograms of alleles observed in either undigested genomic <t>DNA</t> or DNA digested with Hpa II, Hha I or <t>Bst</t> UI from females genotyped via quantitative fluorescent PCR are shown. The boxed numbers correspond to the areas under the allele peaks and the intensity of each peak is in relative fluorescence units (RFU).
    Bst Ui, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 335 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/bst ui/product/New England Biolabs
    Average 95 stars, based on 335 article reviews
    Price from $9.99 to $1999.99
    bst ui - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    95
    New England Biolabs mse i
    Overall strategy for cloning methylated CpG islands. In step 1, genomic <t>DNA</t> was digested with <t>Mse</t> I (red), which cuts between CpG islands, and Hpa II (blue), which cuts unmethylated CpG islands. Mse I fragments containing methylated CpG islands then are transformed into a bacterial strain that does not cut methylated DNA. However, brief bacterial passage leads to loss of methylation of these previously methylated sequences. In step 2, the library DNA is pooled and digested with Eag I (green), which cuts relatively large fragments within CpG islands, and these fragments are then subcloned.
    Mse I, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 1003 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mse i/product/New England Biolabs
    Average 95 stars, based on 1003 article reviews
    Price from $9.99 to $1999.99
    mse i - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    95
    New England Biolabs hin diii
    Overall strategy for cloning methylated CpG islands. In step 1, genomic <t>DNA</t> was digested with <t>Mse</t> I (red), which cuts between CpG islands, and Hpa II (blue), which cuts unmethylated CpG islands. Mse I fragments containing methylated CpG islands then are transformed into a bacterial strain that does not cut methylated DNA. However, brief bacterial passage leads to loss of methylation of these previously methylated sequences. In step 2, the library DNA is pooled and digested with Eag I (green), which cuts relatively large fragments within CpG islands, and these fragments are then subcloned.
    Hin Diii, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 1902 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hin diii/product/New England Biolabs
    Average 95 stars, based on 1902 article reviews
    Price from $9.99 to $1999.99
    hin diii - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    90
    Promega hpa ii
    Methylation-sensitive amplified polymorphism (MSAP) band patterning determined by <t>Hpa</t> II/ <t>Msp</t> I isoschizomers and correspondence with methylation status of CCGG sites. Black squares indicate methylated cytosines.
    Hpa Ii, supplied by Promega, used in various techniques. Bioz Stars score: 90/100, based on 105 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hpa ii/product/Promega
    Average 90 stars, based on 105 article reviews
    Price from $9.99 to $1999.99
    hpa ii - by Bioz Stars, 2020-02
    90/100 stars
      Buy from Supplier

    95
    New England Biolabs hha i
    Methylation statuses at CpG sites near the RP2 onshore tandem GAAA repeat. Random ( A ) and non-random ( B ) X-inactivation patterns generated for different CpG-containing 5 me CpG-sensitive restriction endonuclease sites obtained using the 5 me CpG-based PCR RP2 / AR biplex assay across the restriction sites. Electropherograms of alleles observed in either undigested genomic DNA or DNA digested with <t>Hpa</t> II, <t>Hha</t> I or Bst UI from females genotyped via quantitative fluorescent PCR are shown. The boxed numbers correspond to the areas under the allele peaks and the intensity of each peak is in relative fluorescence units (RFU).
    Hha I, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 584 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hha i/product/New England Biolabs
    Average 95 stars, based on 584 article reviews
    Price from $9.99 to $1999.99
    hha i - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    95
    New England Biolabs xho i
    Restriction fragment length polymorphism analysis of the TcMK amplification product. A: a multiple sequence alignment of the 5′-UTR just upstream of the translational start codon, showing the polymorphic <t>Xho</t> I site. A solid line box marks the Xho I site in TcV strains. B: Agarose gel electrophoresis showing a PCR-RFLP analysis of selected strains. Strains analyzed were: Sc43 cl9, MN cl2, LL014 and Teh53 for DTU TcV; and Tulahuen cl2, CL-Brener, Tul2 and P63 cl1 for DTU TcVI.
    Xho I, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 3877 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/xho i/product/New England Biolabs
    Average 95 stars, based on 3877 article reviews
    Price from $9.99 to $1999.99
    xho i - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    91
    Thermo Fisher hpa ii
    Consensus sequence of the SSUsat monomers from Spisula subtruncata . On the basis of the DNA sequences of the recovered SSUsat monomers from Spisula subtruncata genome, a consensus sequence was derived. Restriction sites for <t>Msp</t> I/ <t>Hpa</t> II, Pvu II and Taq I are underlined. Green and red arrows indicate the positions of PCR primers used for SSUsat amplification in related species.
    Hpa Ii, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 91/100, based on 229 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hpa ii/product/Thermo Fisher
    Average 91 stars, based on 229 article reviews
    Price from $9.99 to $1999.99
    hpa ii - by Bioz Stars, 2020-02
    91/100 stars
      Buy from Supplier

    95
    New England Biolabs dde i
    Consensus sequence of the SSUsat monomers from Spisula subtruncata . On the basis of the DNA sequences of the recovered SSUsat monomers from Spisula subtruncata genome, a consensus sequence was derived. Restriction sites for <t>Msp</t> I/ <t>Hpa</t> II, Pvu II and Taq I are underlined. Green and red arrows indicate the positions of PCR primers used for SSUsat amplification in related species.
    Dde I, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 246 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dde i/product/New England Biolabs
    Average 95 stars, based on 246 article reviews
    Price from $9.99 to $1999.99
    dde i - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    95
    New England Biolabs sau 3ai
    Consensus sequence of the SSUsat monomers from Spisula subtruncata . On the basis of the DNA sequences of the recovered SSUsat monomers from Spisula subtruncata genome, a consensus sequence was derived. Restriction sites for <t>Msp</t> I/ <t>Hpa</t> II, Pvu II and Taq I are underlined. Green and red arrows indicate the positions of PCR primers used for SSUsat amplification in related species.
    Sau 3ai, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 153 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sau 3ai/product/New England Biolabs
    Average 95 stars, based on 153 article reviews
    Price from $9.99 to $1999.99
    sau 3ai - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    95
    New England Biolabs hinp 1i
    Consensus sequence of the SSUsat monomers from Spisula subtruncata . On the basis of the DNA sequences of the recovered SSUsat monomers from Spisula subtruncata genome, a consensus sequence was derived. Restriction sites for <t>Msp</t> I/ <t>Hpa</t> II, Pvu II and Taq I are underlined. Green and red arrows indicate the positions of PCR primers used for SSUsat amplification in related species.
    Hinp 1i, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hinp 1i/product/New England Biolabs
    Average 95 stars, based on 8 article reviews
    Price from $9.99 to $1999.99
    hinp 1i - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    95
    New England Biolabs mbo i
    Consensus sequence of the SSUsat monomers from Spisula subtruncata . On the basis of the DNA sequences of the recovered SSUsat monomers from Spisula subtruncata genome, a consensus sequence was derived. Restriction sites for <t>Msp</t> I/ <t>Hpa</t> II, Pvu II and Taq I are underlined. Green and red arrows indicate the positions of PCR primers used for SSUsat amplification in related species.
    Mbo I, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 217 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mbo i/product/New England Biolabs
    Average 95 stars, based on 217 article reviews
    Price from $9.99 to $1999.99
    mbo i - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    95
    New England Biolabs ecor i
    Representative variation in MSAP profiles. “→” to red arrows represents variation in DNA methylation between diploid and tetraploid plants; “+” represents fragments obtained after digestion with <t>EcoR</t> I or Hpa II/ Msp I; “−” represents fragments not digested by EcoR I or Hpa II/ Msp I; Type I fragments are nonmethylated and were presented in both the H ( EcoR I or Hpa II digest) and M ( EcoR I or Msp I digest) lanes; Type II are fully methylated and only appeared in the M lanes; Type III are hemimethylated and appeared in the H lanes; Type IV were fragments absent from both H and M lanes in diploid but present in either H or M lane of tetraploid, and vice versa.
    Ecor I, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 959 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ecor i/product/New England Biolabs
    Average 95 stars, based on 959 article reviews
    Price from $9.99 to $1999.99
    ecor i - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    95
    New England Biolabs mcrbc
    Comparison of HM-PCR and HM-WGA-PCR results for eleven CGIs . Genomic DNA from peripheral blood leukocytes was digested with Mock (lane 1), <t>Hpa</t> II or Hha I (lane 2), Msp I (lane 3), or <t>McrBC</t> (lane 4). The digested genomic DNA were used for PCR amplification either directly (left; HM-PCR) or after whole-genome-amplification (right; HM-WGA-PCR) using primer pairs for the eight CGIs on chromosome 21 (A~D) and three CGIs on chromosome 11 (E). Here, when Hha I-digested genomic DNA was used in lane 2, 1 ul of distilled water was used in lane 3 in place of Msp I-digested genomic DNA. PCR products were electrophoresed, stained with ethidium bromide, and visualized by UV illumination. Results of bisulfite sequencing are shown for the eight CGIs on chromosome 21 (A~D). Open and closed circles indicate unmethylated and methylated CpG dinucleotides, respectively. Each row of circles represents each sequenced clone of bisufite PCR products.
    Mcrbc, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 479 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mcrbc/product/New England Biolabs
    Average 95 stars, based on 479 article reviews
    Price from $9.99 to $1999.99
    mcrbc - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    95
    New England Biolabs xba i
    Comparison of HM-PCR and HM-WGA-PCR results for eleven CGIs . Genomic DNA from peripheral blood leukocytes was digested with Mock (lane 1), <t>Hpa</t> II or Hha I (lane 2), Msp I (lane 3), or <t>McrBC</t> (lane 4). The digested genomic DNA were used for PCR amplification either directly (left; HM-PCR) or after whole-genome-amplification (right; HM-WGA-PCR) using primer pairs for the eight CGIs on chromosome 21 (A~D) and three CGIs on chromosome 11 (E). Here, when Hha I-digested genomic DNA was used in lane 2, 1 ul of distilled water was used in lane 3 in place of Msp I-digested genomic DNA. PCR products were electrophoresed, stained with ethidium bromide, and visualized by UV illumination. Results of bisulfite sequencing are shown for the eight CGIs on chromosome 21 (A~D). Open and closed circles indicate unmethylated and methylated CpG dinucleotides, respectively. Each row of circles represents each sequenced clone of bisufite PCR products.
    Xba I, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 2655 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/xba i/product/New England Biolabs
    Average 95 stars, based on 2655 article reviews
    Price from $9.99 to $1999.99
    xba i - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    90
    Thermo Fisher ligase buffer
    Comparison of HM-PCR and HM-WGA-PCR results for eleven CGIs . Genomic DNA from peripheral blood leukocytes was digested with Mock (lane 1), <t>Hpa</t> II or Hha I (lane 2), Msp I (lane 3), or <t>McrBC</t> (lane 4). The digested genomic DNA were used for PCR amplification either directly (left; HM-PCR) or after whole-genome-amplification (right; HM-WGA-PCR) using primer pairs for the eight CGIs on chromosome 21 (A~D) and three CGIs on chromosome 11 (E). Here, when Hha I-digested genomic DNA was used in lane 2, 1 ul of distilled water was used in lane 3 in place of Msp I-digested genomic DNA. PCR products were electrophoresed, stained with ethidium bromide, and visualized by UV illumination. Results of bisulfite sequencing are shown for the eight CGIs on chromosome 21 (A~D). Open and closed circles indicate unmethylated and methylated CpG dinucleotides, respectively. Each row of circles represents each sequenced clone of bisufite PCR products.
    Ligase Buffer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 340 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ligase buffer/product/Thermo Fisher
    Average 90 stars, based on 340 article reviews
    Price from $9.99 to $1999.99
    ligase buffer - by Bioz Stars, 2020-02
    90/100 stars
      Buy from Supplier

    95
    New England Biolabs alu i
    Effect of combined DDM1 and siRNA deficiency on 5S rDNA methylation and aberrant transcripts. A) 5S rDNA hypermethylation in ddm1 is siRNA-dependent: Southern blot comparison of <t>Alu</t> I and Hae III-digested genomic <t>DNA</t> isolated from inflorescences of wild type (WT), ddm1 , and double mutant lines nrpd1 ddm1 , rdr2 ddm1 , and dcl3 ddm1 (top panel). The probe is the same as in Figure 2E . Dilutions of the above digests were also assayed by PCR using 5S LT1 primers (bottom panel). Samples to which no restriction enzyme was added are controls (no digest). B) 5S LT1 is silenced by two overlapping processes: RNA samples from inflorescences of WT, ddm1 and the double mutant panel were analyzed by one-step RT-PCR, performed as described in Figure 1C . Control reactions were performed with ACT2 primers; reverse transcriptase was omitted from duplicate 5S LT1 and ACT2 reactions (no RT).
    Alu I, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 605 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/alu i/product/New England Biolabs
    Average 95 stars, based on 605 article reviews
    Price from $9.99 to $1999.99
    alu i - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    Image Search Results


    The Klf1 exon 2 CGI is unmethylated during development. A ) Schematic of the mouse Klf1 gene across exon 2 and the flanking introns shows salient restriction enzyme sites within the region and a 1.1-kbp cDNA probe used for Southern blot. B ) Genomic DNA samples from whole testis between P5 and P45 (lanes 1–7, left and center panels) and P45 spleen (lane 8). DNAs were purified and double-digested with Eco RI and Bam HI restriction enzymes to liberate the 1.85-kb genomic fragment containing the Klf1 exon 2 CGI. Thereafter, the DNA samples were further digested with Hpa II (left panel) or not digested (center panel) and then Southern blotted using the 1.1-kb probe shown in A . In addition, several samples were methylated using Hpa II -methyltransferase then digested with Hpa II (right panel) and Southern blotted. Because of the total number of samples in this experiment, three Southern blots were generated, and the relevant lanes were used to form composite panels. Data for the P5–P15 lanes (break in Southern blots) were obtained from separate Southern blots. Arrowheads in A , locations of Hpa II restriction sites; FL in B , full-length 1.85-kb Eco RI - Bam HI fragments that are visible in the center and right panels; Spl, spleen.

    Journal: Biology of Reproduction

    Article Title: Tissue-Restricted Transcription from a Conserved Intragenic CpG Island in the Klf1 Gene in Mice 1

    doi: 10.1095/biolreprod.112.099879

    Figure Lengend Snippet: The Klf1 exon 2 CGI is unmethylated during development. A ) Schematic of the mouse Klf1 gene across exon 2 and the flanking introns shows salient restriction enzyme sites within the region and a 1.1-kbp cDNA probe used for Southern blot. B ) Genomic DNA samples from whole testis between P5 and P45 (lanes 1–7, left and center panels) and P45 spleen (lane 8). DNAs were purified and double-digested with Eco RI and Bam HI restriction enzymes to liberate the 1.85-kb genomic fragment containing the Klf1 exon 2 CGI. Thereafter, the DNA samples were further digested with Hpa II (left panel) or not digested (center panel) and then Southern blotted using the 1.1-kb probe shown in A . In addition, several samples were methylated using Hpa II -methyltransferase then digested with Hpa II (right panel) and Southern blotted. Because of the total number of samples in this experiment, three Southern blots were generated, and the relevant lanes were used to form composite panels. Data for the P5–P15 lanes (break in Southern blots) were obtained from separate Southern blots. Arrowheads in A , locations of Hpa II restriction sites; FL in B , full-length 1.85-kb Eco RI - Bam HI fragments that are visible in the center and right panels; Spl, spleen.

    Article Snippet: Genomic DNAs purified from testes were restriction digested with Eco RI and Bam HI (New England Biolabs) at 37°C for 4–6 h. DNA samples were then either restriction digested with Hpa II (New England Biolabs) for 4–6 h or methylated in vitro using Hpa II methyltransferase (New England Biolabs) and digested with Hpa II .

    Techniques: Southern Blot, Purification, Methylation, Generated

    Methylation analysis of the ZPBP gene in whole blood samples of Berkshire sows by PMP assay. gDNAs were cut with the methylation-sensitive restriction enzymes Hpa II/ Msp I. M: size marker; U: undigested DNA; SLG: smaller litter size group; LLG: larger litter size group.

    Journal: Archives Animal Breeding

    Article Title: Hypomethylation in the promoter region of ZPBP as a potential litter size indicator in Berkshire pigs

    doi: 10.5194/aab-62-69-2019

    Figure Lengend Snippet: Methylation analysis of the ZPBP gene in whole blood samples of Berkshire sows by PMP assay. gDNAs were cut with the methylation-sensitive restriction enzymes Hpa II/ Msp I. M: size marker; U: undigested DNA; SLG: smaller litter size group; LLG: larger litter size group.

    Article Snippet: To verify the result of GWBS, whole blood samples were collected from Berkshire sows to isolate gDNA using a Wizard Genomic DNA Purification Kit (Promega, Madison, WI, USA) according to the manufacturer's instructions and digested with Hpa II (NEB, Hitchin, UK) and Msp I (NEB), a pair of methylation-sensitive isoschizomers that have the same recognition site (CC|GG).

    Techniques: Methylation, Marker

    pRF/FMDV-IRES and pCAGGS/FMDV-IRES plasmid construction. Structure of the bicistronic luciferase reporter construct containing the FMDV-IRES element located between Renilla luciferase and firefly luciferase (pRF-FMDV-IRS). Reporter gene was excised from this plasmid construct using the restriction enzymes Eco RV and Hpa I, and was o the pCAGGS/MCS(F) vector treated with Sma I and rAPid Alkaline Phosphatase using Mighty Mix

    Journal: BMC Veterinary Research

    Article Title: IRES-mediated translation of foot-and-mouth disease virus (FMDV) in cultured cells derived from FMDV-susceptible and -insusceptible animals

    doi: 10.1186/s12917-016-0694-8

    Figure Lengend Snippet: pRF/FMDV-IRES and pCAGGS/FMDV-IRES plasmid construction. Structure of the bicistronic luciferase reporter construct containing the FMDV-IRES element located between Renilla luciferase and firefly luciferase (pRF-FMDV-IRS). Reporter gene was excised from this plasmid construct using the restriction enzymes Eco RV and Hpa I, and was o the pCAGGS/MCS(F) vector treated with Sma I and rAPid Alkaline Phosphatase using Mighty Mix

    Article Snippet: Reporter genes were excised from pRF/FMDV-IRES using the restriction endonucleases Eco RV (Toyobo) and Hpa I (NEB). pCAGGS/FMDV-IRES was generated by inserting a reporter gene into pCAGGS/MSC(F), which was then treated with SmaI (Takara) and rAPid Alkaline Phosphatase (Roche) using Mighty Mix (Takara).

    Techniques: Plasmid Preparation, Luciferase, Construct

    Examples of MSAP profiles showing various types of locus-specific DNA methylation inheritance and variation in F 1 hybrid and their parents (a) and different types of locus-specific DNA methylation inheritance and variation in F 4 and F 10 , and the F 4 and F 10 DNA methylation patterns (b,c) . (a). Primer combination is Eco RI+ ATT/ Hpa II ( Msp I) + ACC. Amplification results of Hpa II-digested genomic DNA of R. sativus L. (Lane 1), B. alboglabra Bailey (Lane 3) and F 1 hybrids (Lane 5) Amplification results of Msp I-digested genomic DNA of R. sativus L. (Lane 2), B. alboglabra Bailey (Lane 4), and F 1 hybrids (Lane 6). (b) and (c). Primer combinations are Eco RI + AGG/ Hpa II ( Msp I) + GAT and Eco RI + AGG/ Hpa II ( Msp I) + TCG, respectively. Arrows show variations of the various methylation patterns. H: Hpa II, M: Msp I.

    Journal: BMC Plant Biology

    Article Title: Instability of chromosome number and DNA methylation variation induced by hybridization and amphidiploid formation between Raphanus sativus L. and Brassica alboglabra Bailey

    doi: 10.1186/1471-2229-10-207

    Figure Lengend Snippet: Examples of MSAP profiles showing various types of locus-specific DNA methylation inheritance and variation in F 1 hybrid and their parents (a) and different types of locus-specific DNA methylation inheritance and variation in F 4 and F 10 , and the F 4 and F 10 DNA methylation patterns (b,c) . (a). Primer combination is Eco RI+ ATT/ Hpa II ( Msp I) + ACC. Amplification results of Hpa II-digested genomic DNA of R. sativus L. (Lane 1), B. alboglabra Bailey (Lane 3) and F 1 hybrids (Lane 5) Amplification results of Msp I-digested genomic DNA of R. sativus L. (Lane 2), B. alboglabra Bailey (Lane 4), and F 1 hybrids (Lane 6). (b) and (c). Primer combinations are Eco RI + AGG/ Hpa II ( Msp I) + GAT and Eco RI + AGG/ Hpa II ( Msp I) + TCG, respectively. Arrows show variations of the various methylation patterns. H: Hpa II, M: Msp I.

    Article Snippet: The restriction enzymes Eco RI, Hpa II, and Msp I were purchased from the New England Biolabs Inc. (Beverly, Mass.).

    Techniques: DNA Methylation Assay, Amplification, Methylation

    Methylation states of the 5′ regions of the leukosialin gene exogenously introduced and stably maintained in non-leukosialin-expressing HeLa cells. (A) Schematic representations of the leukosialin gene and leukosialin CAT constructs. The exons are depicted by filled boxes, and introns and the 5′ flanking regions are depicted with horizontal lines. The vector sequences are depicted with dotted lines. Msp I (CCGG) sites are shown with vertical bars. The asterisk indicates the polymorphic site, where an Msp I site is lost in one allele of HeLa cells. The Msp I DNA fragment (560 bp) used for a hybridization probe is presented at the top. (B) Genomic DNAs (10 μg) from HeLa cells stably transfected with CpG-methylated- or unmethylated-leukosialin CAT constructs were digested with Hpa II (H) or Msp I (M) and separated by 1.5% agarose gel electrophoresis. The blotted filter was hybridized with the 560-bp DNA fragment of the 5′ region of the leukosialin gene shown in panel A. The hatched arrow indicates the position of the fragments produced by the endogenous gene as well as the exogenously introduced gene. The filled arrow indicates the signal produced by the exogenously introduced gene. The open arrow indicates the position of a polymorphic fragment of the endogenous leukosialin gene of HeLa cells.

    Journal: Molecular and Cellular Biology

    Article Title: Methyl-CpG-Binding Protein MeCP2 Represses Sp1-Activated Transcription of the Human Leukosialin Gene When the Promoter Is Methylated

    doi:

    Figure Lengend Snippet: Methylation states of the 5′ regions of the leukosialin gene exogenously introduced and stably maintained in non-leukosialin-expressing HeLa cells. (A) Schematic representations of the leukosialin gene and leukosialin CAT constructs. The exons are depicted by filled boxes, and introns and the 5′ flanking regions are depicted with horizontal lines. The vector sequences are depicted with dotted lines. Msp I (CCGG) sites are shown with vertical bars. The asterisk indicates the polymorphic site, where an Msp I site is lost in one allele of HeLa cells. The Msp I DNA fragment (560 bp) used for a hybridization probe is presented at the top. (B) Genomic DNAs (10 μg) from HeLa cells stably transfected with CpG-methylated- or unmethylated-leukosialin CAT constructs were digested with Hpa II (H) or Msp I (M) and separated by 1.5% agarose gel electrophoresis. The blotted filter was hybridized with the 560-bp DNA fragment of the 5′ region of the leukosialin gene shown in panel A. The hatched arrow indicates the position of the fragments produced by the endogenous gene as well as the exogenously introduced gene. The filled arrow indicates the signal produced by the exogenously introduced gene. The open arrow indicates the position of a polymorphic fragment of the endogenous leukosialin gene of HeLa cells.

    Article Snippet: These CAT plasmid DNAs were treated with Sss I (CpG) methylase or Hpa II methylase (New England Biolabs) in the presence (methylated) or absence (unmethylated) of 5 mM S -adenosylmethionine.

    Techniques: Methylation, Stable Transfection, Expressing, Construct, Plasmid Preparation, Hybridization, Transfection, Agarose Gel Electrophoresis, Produced

    Transactivation of methylated-leukosialin promoter activity by Sp1 in Drosophila cells. The CAT constructs were methylated in vitro with Hpa II or Sss I (CpG) methylase. Methylated or unmethylated CAT constructs (4 μg) were transfected with 0.5 μg of A5C (filled bars) or with 0.5 μg of pPacSp1 (hatched bars) into Drosophila SL2 cells. After 48 h, the cellular extract was subjected to the CAT assay. CAT activity is presented as a percentage of the level of conversion of the acetylated form. Each value is the mean and standard deviation of results from three independent experiments.

    Journal: Molecular and Cellular Biology

    Article Title: Methyl-CpG-Binding Protein MeCP2 Represses Sp1-Activated Transcription of the Human Leukosialin Gene When the Promoter Is Methylated

    doi:

    Figure Lengend Snippet: Transactivation of methylated-leukosialin promoter activity by Sp1 in Drosophila cells. The CAT constructs were methylated in vitro with Hpa II or Sss I (CpG) methylase. Methylated or unmethylated CAT constructs (4 μg) were transfected with 0.5 μg of A5C (filled bars) or with 0.5 μg of pPacSp1 (hatched bars) into Drosophila SL2 cells. After 48 h, the cellular extract was subjected to the CAT assay. CAT activity is presented as a percentage of the level of conversion of the acetylated form. Each value is the mean and standard deviation of results from three independent experiments.

    Article Snippet: These CAT plasmid DNAs were treated with Sss I (CpG) methylase or Hpa II methylase (New England Biolabs) in the presence (methylated) or absence (unmethylated) of 5 mM S -adenosylmethionine.

    Techniques: Methylation, Activity Assay, Construct, In Vitro, Transfection, Standard Deviation

    DNMT3L stimulates de novo methylation by Dnmt3a but not by DNMT3B at the SNRPN IC. DNA harvested from human 293/EBNA1 cells carrying pFC19 or pFC30 (in which a key ≈1-kb region of the SNRPN IC was cloned in either orientation) was digested with an excess of the methylation-sensitive enzyme Hpa II. The resulting DNA species were separated by electrophoresis through an agarose gel, transferred to a nylon membrane, and hybridized with a probe corresponding to the SNRPN IC. Two bands (684 and 754 bp for pFC19, and 684 and 221 bp for pFC30) were expected if no methylation was present. CpG methylation prevents cleavage by Hpa II and results in higher molecular weight species that can be easily separated. ( A ) Each target plasmid was introduced into cells alone, together with an expression vector for Dnmt3a, or together with expression vectors for both Dnmt3a and DNMT3L, as indicated. ( B ) Plasmid pFC19 was introduced in cells either with DNMT3B or with both DNMT3B and DNMT3L expression vectors, as indicated.

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

    Article Title: The DNA methyltransferase-like protein DNMT3L stimulates de novo methylation by Dnmt3a

    doi: 10.1073/pnas.262443999

    Figure Lengend Snippet: DNMT3L stimulates de novo methylation by Dnmt3a but not by DNMT3B at the SNRPN IC. DNA harvested from human 293/EBNA1 cells carrying pFC19 or pFC30 (in which a key ≈1-kb region of the SNRPN IC was cloned in either orientation) was digested with an excess of the methylation-sensitive enzyme Hpa II. The resulting DNA species were separated by electrophoresis through an agarose gel, transferred to a nylon membrane, and hybridized with a probe corresponding to the SNRPN IC. Two bands (684 and 754 bp for pFC19, and 684 and 221 bp for pFC30) were expected if no methylation was present. CpG methylation prevents cleavage by Hpa II and results in higher molecular weight species that can be easily separated. ( A ) Each target plasmid was introduced into cells alone, together with an expression vector for Dnmt3a, or together with expression vectors for both Dnmt3a and DNMT3L, as indicated. ( B ) Plasmid pFC19 was introduced in cells either with DNMT3B or with both DNMT3B and DNMT3L expression vectors, as indicated.

    Article Snippet: The recovered DNA was digested overnight with an excess of Hpa II enzyme (New England Biolabs).

    Techniques: Methylation, Clone Assay, Electrophoresis, Agarose Gel Electrophoresis, CpG Methylation Assay, Molecular Weight, Plasmid Preparation, Expressing

    ) were incubated the methylation-sensitive restriction enzyme Hpa II, which cleaves only unmethylated DNA; the PCR products then were subjected to capillary gel electrophoresis. The boxed numbers represent the area under each peak (ar), the peak height (ht), and the length (sz) of the DNA fragment. Note that peak area and height for the PCR products derived from individual S2-II/6 are reduced in comparison to the amplicon derived from S2-II/5, indicating a partial loss of methylation.

    Journal: Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research

    Article Title: Exclusion of the GNAS Locus in PHP-Ib Patients With Broad GNAS Methylation Changes: Evidence for an Autosomal Recessive Form of PHP-Ib?

    doi: 10.1002/jbmr.408

    Figure Lengend Snippet: ) were incubated the methylation-sensitive restriction enzyme Hpa II, which cleaves only unmethylated DNA; the PCR products then were subjected to capillary gel electrophoresis. The boxed numbers represent the area under each peak (ar), the peak height (ht), and the length (sz) of the DNA fragment. Note that peak area and height for the PCR products derived from individual S2-II/6 are reduced in comparison to the amplicon derived from S2-II/5, indicating a partial loss of methylation.

    Article Snippet: Genomic DNA from a patUPD20q patient, a healthy individual from family 3 (S2-II/5) and two affected family members, S2-II/3 and S2-II/6, were incubated with the restriction enzyme Hpa II (New England Biolabs, Ipswich, MA, USA), which cleaves only the unmethylated allele within the A/B region.

    Techniques: Incubation, Methylation, Polymerase Chain Reaction, Nucleic Acid Electrophoresis, Derivative Assay, Amplification

    Representative examples of the electrophoretic pattern. ( A ) Male control (digestion control; polymerase chain reaction [PCR] products from undigested DNA appear as a single band, while PCR products from digested DNA does not have bands, this shows DNA is completely digested with Hpa II). ( B ) Excluded sample (could not distinguish two bands of undigested PCR products because two alleles from the maternally and paternally inherited CAG repeats were the same). ( C ) Not skewed (degree of X chromosome inactivation [XCI] skewing: 55.3%). ( D ) Skewed (degree of XCI skewing: 88.2%). Lane: + and − indicate undigestioned and digestion by Hpa II, respectively.

    Journal: Thyroid

    Article Title: The Relationship Between Skewed X Chromosome Inactivation and the Prognosis of Graves' and Hashimoto's Diseases

    doi: 10.1089/thy.2014.0318

    Figure Lengend Snippet: Representative examples of the electrophoretic pattern. ( A ) Male control (digestion control; polymerase chain reaction [PCR] products from undigested DNA appear as a single band, while PCR products from digested DNA does not have bands, this shows DNA is completely digested with Hpa II). ( B ) Excluded sample (could not distinguish two bands of undigested PCR products because two alleles from the maternally and paternally inherited CAG repeats were the same). ( C ) Not skewed (degree of X chromosome inactivation [XCI] skewing: 55.3%). ( D ) Skewed (degree of XCI skewing: 88.2%). Lane: + and − indicate undigestioned and digestion by Hpa II, respectively.

    Article Snippet: Isolated DNA was digested with the methylation sensitive restriction enzyme Hpa II (New England BioLabs, Inc., Beverly, NA) to obtain polymerase chain reaction (PCR) templates.

    Techniques: Polymerase Chain Reaction

    Detailed MSAP-Seq assay overview (A) Genomic DNA is cleaved using rare cutter ( Eco RI) and methylation sensitive restriction enzyme ( Hpa II); only unmethylated recognition sites are digested by Hpa II. Then adapters specific to sticky ends are ligated and obtained fragments are amplified in PCR using selective primers; only fragments generated from unmethylated regions are amplified as they contain ends complementary to adapters. Products are then purified and fragmented by sonication to create shorter tags. (B) Purified fragments are used for standard library preparation involving following steps: end repair, adenylation, barcoded adapters ligation and purification, PCR amplification and purification. Then libraries quality and quantity is estimated. (C) Prepared libraries are pooled and processed thru cluster generation and high-throughput sequencing. (D) Sequencing data are analyzed using dedicated automatic pipeline—MSEQER. Firstly, reads are filtered for presence of Hpa II adapter and adapters are clipped. Then only reads containing CGG tags on the ends are mapped to the reference genome and functionally annotated. Obtained counts at each of the CCGG sites are normalized and differential methylation analysis among sets of samples is performed.

    Journal: Frontiers in Plant Science

    Article Title: Methylation Sensitive Amplification Polymorphism Sequencing (MSAP-Seq)—A Method for High-Throughput Analysis of Differentially Methylated CCGG Sites in Plants with Large Genomes

    doi: 10.3389/fpls.2017.02056

    Figure Lengend Snippet: Detailed MSAP-Seq assay overview (A) Genomic DNA is cleaved using rare cutter ( Eco RI) and methylation sensitive restriction enzyme ( Hpa II); only unmethylated recognition sites are digested by Hpa II. Then adapters specific to sticky ends are ligated and obtained fragments are amplified in PCR using selective primers; only fragments generated from unmethylated regions are amplified as they contain ends complementary to adapters. Products are then purified and fragmented by sonication to create shorter tags. (B) Purified fragments are used for standard library preparation involving following steps: end repair, adenylation, barcoded adapters ligation and purification, PCR amplification and purification. Then libraries quality and quantity is estimated. (C) Prepared libraries are pooled and processed thru cluster generation and high-throughput sequencing. (D) Sequencing data are analyzed using dedicated automatic pipeline—MSEQER. Firstly, reads are filtered for presence of Hpa II adapter and adapters are clipped. Then only reads containing CGG tags on the ends are mapped to the reference genome and functionally annotated. Obtained counts at each of the CCGG sites are normalized and differential methylation analysis among sets of samples is performed.

    Article Snippet: Identification of differentially methylated sites (DMS) using MSAP-Seq Five hundred nanograms of genomic DNA was cut with 2.5 U of the frequent-cutting methylation sensitive restriction enzyme Hpa II (New England Biolabs, Ipswich, USA) and 2.5 U of the rare-cutting Eco RI (New England Biolabs, Ipswich, USA) in a 20 μL reaction with 1x NEB1 buffer (New England Biolabs, Ipswich, USA) at 37°C for 6 h. Enzymes were inactivated at 80°C for 20 min. Next, 12 μL of ligation mixture containing 60 pmol of Hpa II-related adapter, 6 pmol of Eco RI-related adapter (Table ), 1x T4 ligase buffer (Thermo Scientific, Waltham, USA) and 1.2 U of T4 DNA ligase (Thermo Scientific, Waltham, USA) were added.

    Techniques: Methylation, Amplification, Polymerase Chain Reaction, Generated, Purification, Sonication, Ligation, Next-Generation Sequencing, Sequencing

    Flowchart of MSD-library preparation. Genomic DNA (100 ng) was digested with 10 units of the primary restriction enzyme Sbf I for 1 h and then ligated with 0.5 nmol Adaptor A using 400 units of T4 DNA ligase for 2 h. The treated sample was then digested with 100 units of the non-methylation-sensitive restriction enzyme Msp I (100 units) followed by ligation of the ends of the DNA fragment with Adaptor B. The ligated DNA fragments were then digested with 50 units of Hpa II for 1 h. Owing to the methylation sensitivity of Hap II, only DNA fragments with a methylated CpG retained Adaptor B, which was removed from all other fragments. The DNA fragments were then subjected to Pre-PCR using specific primers for Adaptor A and Adaptor B. Fragments that did not contain Adaptor B at this stage were not amplified. The Pre-PCR amplicons (MSD library) were then amplified as a subpopulation by selective-PCR with 6-carboxyfluorescein (6-FAM)-labelled selective-PCR primers. Finally, the selective-PCR products were electrophoresed with a capillary sequencer and separated by length

    Journal: BMC Molecular Biology

    Article Title: Methylated site display (MSD)-AFLP, a sensitive and affordable method for analysis of CpG methylation profiles

    doi: 10.1186/s12867-017-0083-2

    Figure Lengend Snippet: Flowchart of MSD-library preparation. Genomic DNA (100 ng) was digested with 10 units of the primary restriction enzyme Sbf I for 1 h and then ligated with 0.5 nmol Adaptor A using 400 units of T4 DNA ligase for 2 h. The treated sample was then digested with 100 units of the non-methylation-sensitive restriction enzyme Msp I (100 units) followed by ligation of the ends of the DNA fragment with Adaptor B. The ligated DNA fragments were then digested with 50 units of Hpa II for 1 h. Owing to the methylation sensitivity of Hap II, only DNA fragments with a methylated CpG retained Adaptor B, which was removed from all other fragments. The DNA fragments were then subjected to Pre-PCR using specific primers for Adaptor A and Adaptor B. Fragments that did not contain Adaptor B at this stage were not amplified. The Pre-PCR amplicons (MSD library) were then amplified as a subpopulation by selective-PCR with 6-carboxyfluorescein (6-FAM)-labelled selective-PCR primers. Finally, the selective-PCR products were electrophoresed with a capillary sequencer and separated by length

    Article Snippet: Reagents The reagents and materials used in this study were purchased from the manufacturers indicated in parentheses: CpG methyltransferase (M.Sss I), T4 DNA ligase, and restriction enzymes Hpa II, Msp I, Sbf I, and Stu I (New England Biolabs, MA, USA) it guarantees that the efficiency of their restriction enzymes is almost and the methylation of CpG blocks 100% Hpa II digestion reaction; EpiTect Bisulfite Kit and AllPrep DNA/RNA Mini Kit (Qiagen, Hilden, Germany); Oligonucleotides (Operon, Alameda, CA, USA); Magnetic beads coated with streptavidin (Dynabeads® M-280 Streptavidin) (Dynal, Oslo, Norway); TITANIUM Taq DNA polymerase (Takara Bio, Kusatsu, Japan); GenElute™ Agarose Spin Columns (Sigma-Aldrich, St. Louis, MO, USA); Ligation Convenience Kit (Nippon Gene, Tokyo, Japan); pGEM® -T Easy Vector (Promega, Madison, WI, USA); Competent Cell DH5α and Insert Check-Ready (Toyobo, Osaka, Japan); LightCycler® 480 SYBR Green I Master (Roche Diagnostics GmbH, Mannheim, Germany); POP-7™ Polymer, GeneScan™ 500 LIZ® Size Standard, and BigDye® Terminator v3.1 Cycle Sequencing Kit (ThermoFisher Scientific Inc., San Diego, CA, USA).

    Techniques: Methylation, Ligation, Polymerase Chain Reaction, Amplification

    Resistance of methylated B. bovis DNA to cleavage by Hpa II. DNAs prepared from E. coli or B. bovis were treated with Sss I methylase and compared with untreated DNA for sensitivity to Hpa II digestion. DNA was visualized after electrophoresis on ethidium bromide-stained agarose gels. Lanes: M, 1-kb markers; 1, untreated DNA; 2, untreated DNA incubated with Hpa II; 3, methylated DNA; 4, methylated DNA incubated with Hpa II.

    Journal: Infection and Immunity

    Article Title: DNA and a CpG Oligonucleotide Derived from Babesia bovis Are Mitogenic for Bovine B Cells

    doi:

    Figure Lengend Snippet: Resistance of methylated B. bovis DNA to cleavage by Hpa II. DNAs prepared from E. coli or B. bovis were treated with Sss I methylase and compared with untreated DNA for sensitivity to Hpa II digestion. DNA was visualized after electrophoresis on ethidium bromide-stained agarose gels. Lanes: M, 1-kb markers; 1, untreated DNA; 2, untreated DNA incubated with Hpa II; 3, methylated DNA; 4, methylated DNA incubated with Hpa II.

    Article Snippet: CpG methylase ( Sss I methylase) and Hpa II restriction endonuclease were purchased from New England BioLabs (Beverly, Mass.), and methylation was performed according to the manufacturer’s instructions.

    Techniques: Methylation, Electrophoresis, Staining, Incubation

    Identification of 669insA in exon 4 of Gng3lg . A, DNA sequence analysis of subject CGL-F1. Sequencing of the opposite strand showing the insertion of a T is shown. The subject was homozygous for the insertion. B, PCR-RFLP of 669insA in affected individuals and family members. The insertion, 669insA, introduces a Hpa I restriction site. Lane 1, Fifty-base-pair DNA ladder; lane 2, homozygous normal (unaffected); lane 4, homozygous for 669insA (affected); lanes 3 and 5–8, heterozygous for 669insA (unaffected).

    Journal: The Journal of Clinical Endocrinology and Metabolism

    Article Title: Mutations in Gng3lg and AGPAT2 in Berardinelli-Seip Congenital Lipodystrophy and Brunzell Syndrome: Phenotype Variability Suggests Important Modifier Effects

    doi: 10.1210/jc.2003-030485

    Figure Lengend Snippet: Identification of 669insA in exon 4 of Gng3lg . A, DNA sequence analysis of subject CGL-F1. Sequencing of the opposite strand showing the insertion of a T is shown. The subject was homozygous for the insertion. B, PCR-RFLP of 669insA in affected individuals and family members. The insertion, 669insA, introduces a Hpa I restriction site. Lane 1, Fifty-base-pair DNA ladder; lane 2, homozygous normal (unaffected); lane 4, homozygous for 669insA (affected); lanes 3 and 5–8, heterozygous for 669insA (unaffected).

    Article Snippet: Five microliters of exon 4 PCR product, which were generated with upstream primer 5′-TTGTGTGTCAAGGGTCCTCA-3′ and downstream primer 5′-AAAACAAGACCCCCACATCA-3′, were digested with 7.5 U of Hpa I restriction endonuclease (New England BioLabs, Beverly, MA) for 3 h at 37 C. The fragments were separated on a 2% agarose gel and visualized after staining with ethidium bromide.

    Techniques: Sequencing, Polymerase Chain Reaction

    Genetic characterization Trypanosoma cruzi and Trypanosoma rangeli strains. (A) kDNA analysis of T . rangeli strains in a silver stained 6% polyacrylamide gel containing PCR products obtained with primers S35/S36/KP1L. The presence of a 165-bp band indicates the presence of KP1 minicircles. (B) Detection of the 832-bp fragment of TcSC5D gene in T . cruzi strains and clone. (C) PCR–RFLP of TcSC5D products digested with Hpa I and Sph I enzymes. MM: Molecular marker 100bp. NTC: No-template control.

    Journal: PLoS ONE

    Article Title: DNA content analysis allows discrimination between Trypanosoma cruzi and Trypanosoma rangeli

    doi: 10.1371/journal.pone.0189907

    Figure Lengend Snippet: Genetic characterization Trypanosoma cruzi and Trypanosoma rangeli strains. (A) kDNA analysis of T . rangeli strains in a silver stained 6% polyacrylamide gel containing PCR products obtained with primers S35/S36/KP1L. The presence of a 165-bp band indicates the presence of KP1 minicircles. (B) Detection of the 832-bp fragment of TcSC5D gene in T . cruzi strains and clone. (C) PCR–RFLP of TcSC5D products digested with Hpa I and Sph I enzymes. MM: Molecular marker 100bp. NTC: No-template control.

    Article Snippet: Aliquots of 20 μL of the amplified products were digested with 1U of the enzyme Hpa I (NEB R105) at 55°C for 1 h and with 1U of the enzyme Sph I (NEB R0182) at 37°C for 1 h (TcSC5D fragment) or with one unit of Xho I (NEB R0146S) endonuclease (TcMK fragment).

    Techniques: Staining, Polymerase Chain Reaction, Marker

    Methylation statuses at CpG sites near the RP2 onshore tandem GAAA repeat. Random ( A ) and non-random ( B ) X-inactivation patterns generated for different CpG-containing 5 me CpG-sensitive restriction endonuclease sites obtained using the 5 me CpG-based PCR RP2 / AR biplex assay across the restriction sites. Electropherograms of alleles observed in either undigested genomic DNA or DNA digested with Hpa II, Hha I or Bst UI from females genotyped via quantitative fluorescent PCR are shown. The boxed numbers correspond to the areas under the allele peaks and the intensity of each peak is in relative fluorescence units (RFU).

    Journal: PLoS ONE

    Article Title: 5meCpG Epigenetic Marks Neighboring a Primate-Conserved Core Promoter Short Tandem Repeat Indicate X-Chromosome Inactivation

    doi: 10.1371/journal.pone.0103714

    Figure Lengend Snippet: Methylation statuses at CpG sites near the RP2 onshore tandem GAAA repeat. Random ( A ) and non-random ( B ) X-inactivation patterns generated for different CpG-containing 5 me CpG-sensitive restriction endonuclease sites obtained using the 5 me CpG-based PCR RP2 / AR biplex assay across the restriction sites. Electropherograms of alleles observed in either undigested genomic DNA or DNA digested with Hpa II, Hha I or Bst UI from females genotyped via quantitative fluorescent PCR are shown. The boxed numbers correspond to the areas under the allele peaks and the intensity of each peak is in relative fluorescence units (RFU).

    Article Snippet: Digestion with methylation-sensitive restriction enzymes Genomic DNA (500 ng) was digested with Hpa II (Invitrogen, Carlsbad, CA, USA), Bst UI and Hha I (New England Biolabs, Ipswich, MA, USA) for 6 h at 37°C (Hpa II and Hha I) or 60°C (Bst UI), or was mock-digested without the restriction enzymes.

    Techniques: Methylation, Generated, Polymerase Chain Reaction, Fluorescence

    Overall strategy for cloning methylated CpG islands. In step 1, genomic DNA was digested with Mse I (red), which cuts between CpG islands, and Hpa II (blue), which cuts unmethylated CpG islands. Mse I fragments containing methylated CpG islands then are transformed into a bacterial strain that does not cut methylated DNA. However, brief bacterial passage leads to loss of methylation of these previously methylated sequences. In step 2, the library DNA is pooled and digested with Eag I (green), which cuts relatively large fragments within CpG islands, and these fragments are then subcloned.

    Journal: Genome Research

    Article Title: A Genome-Wide Screen for Normally Methylated Human CpG Islands That Can Identify Novel Imprinted Genes

    doi: 10.1101/gr.224102

    Figure Lengend Snippet: Overall strategy for cloning methylated CpG islands. In step 1, genomic DNA was digested with Mse I (red), which cuts between CpG islands, and Hpa II (blue), which cuts unmethylated CpG islands. Mse I fragments containing methylated CpG islands then are transformed into a bacterial strain that does not cut methylated DNA. However, brief bacterial passage leads to loss of methylation of these previously methylated sequences. In step 2, the library DNA is pooled and digested with Eag I (green), which cuts relatively large fragments within CpG islands, and these fragments are then subcloned.

    Article Snippet: Genomic DNA was digested with Mse I alone or Mse I together with a methylcytosine-sensitive (Hpa II, LTI, or Sma I, NEB) or methyl-insensitive (Msp I or Xma I, NEB) restriction endonuclease according to the manufacturer’s conditions.

    Techniques: Clone Assay, Methylation, Transformation Assay

    Methylation-sensitive amplified polymorphism (MSAP) band patterning determined by Hpa II/ Msp I isoschizomers and correspondence with methylation status of CCGG sites. Black squares indicate methylated cytosines.

    Journal: Ecology and Evolution

    Article Title: Epigenetic patterns newly established after interspecific hybridization in natural populations of Solanum

    doi: 10.1002/ece3.758

    Figure Lengend Snippet: Methylation-sensitive amplified polymorphism (MSAP) band patterning determined by Hpa II/ Msp I isoschizomers and correspondence with methylation status of CCGG sites. Black squares indicate methylated cytosines.

    Article Snippet: The digest was split into two equal volumes and 10 U of Hpa II (Promega) and 10 U of Msp I (New England Biolabs) were added to each half, along with 100 ng·μL−1 BSA and the appropriate buffers in a final volume of 20 μL for the second digestion.

    Techniques: Methylation, Amplification

    Methylation statuses at CpG sites near the RP2 onshore tandem GAAA repeat. Random ( A ) and non-random ( B ) X-inactivation patterns generated for different CpG-containing 5 me CpG-sensitive restriction endonuclease sites obtained using the 5 me CpG-based PCR RP2 / AR biplex assay across the restriction sites. Electropherograms of alleles observed in either undigested genomic DNA or DNA digested with Hpa II, Hha I or Bst UI from females genotyped via quantitative fluorescent PCR are shown. The boxed numbers correspond to the areas under the allele peaks and the intensity of each peak is in relative fluorescence units (RFU).

    Journal: PLoS ONE

    Article Title: 5meCpG Epigenetic Marks Neighboring a Primate-Conserved Core Promoter Short Tandem Repeat Indicate X-Chromosome Inactivation

    doi: 10.1371/journal.pone.0103714

    Figure Lengend Snippet: Methylation statuses at CpG sites near the RP2 onshore tandem GAAA repeat. Random ( A ) and non-random ( B ) X-inactivation patterns generated for different CpG-containing 5 me CpG-sensitive restriction endonuclease sites obtained using the 5 me CpG-based PCR RP2 / AR biplex assay across the restriction sites. Electropherograms of alleles observed in either undigested genomic DNA or DNA digested with Hpa II, Hha I or Bst UI from females genotyped via quantitative fluorescent PCR are shown. The boxed numbers correspond to the areas under the allele peaks and the intensity of each peak is in relative fluorescence units (RFU).

    Article Snippet: Digestion with methylation-sensitive restriction enzymes Genomic DNA (500 ng) was digested with Hpa II (Invitrogen, Carlsbad, CA, USA), Bst UI and Hha I (New England Biolabs, Ipswich, MA, USA) for 6 h at 37°C (Hpa II and Hha I) or 60°C (Bst UI), or was mock-digested without the restriction enzymes.

    Techniques: Methylation, Generated, Polymerase Chain Reaction, Fluorescence

    Restriction fragment length polymorphism analysis of the TcMK amplification product. A: a multiple sequence alignment of the 5′-UTR just upstream of the translational start codon, showing the polymorphic Xho I site. A solid line box marks the Xho I site in TcV strains. B: Agarose gel electrophoresis showing a PCR-RFLP analysis of selected strains. Strains analyzed were: Sc43 cl9, MN cl2, LL014 and Teh53 for DTU TcV; and Tulahuen cl2, CL-Brener, Tul2 and P63 cl1 for DTU TcVI.

    Journal: PLoS Neglected Tropical Diseases

    Article Title: A Simple Strain Typing Assay for Trypanosoma cruzi: Discrimination of Major Evolutionary Lineages from a Single Amplification Product

    doi: 10.1371/journal.pntd.0001777

    Figure Lengend Snippet: Restriction fragment length polymorphism analysis of the TcMK amplification product. A: a multiple sequence alignment of the 5′-UTR just upstream of the translational start codon, showing the polymorphic Xho I site. A solid line box marks the Xho I site in TcV strains. B: Agarose gel electrophoresis showing a PCR-RFLP analysis of selected strains. Strains analyzed were: Sc43 cl9, MN cl2, LL014 and Teh53 for DTU TcV; and Tulahuen cl2, CL-Brener, Tul2 and P63 cl1 for DTU TcVI.

    Article Snippet: The resulting restriction fragments were resolved by electrophoresis of in 2% TBE-agarose gels, applying 5,33 V/cm for 1 h. For TcMK fragment, an aliquot (20 µl) of the amplification products where digested in a single incubation with 1 U of Xho I (NEB R0146) at 37 for 1 h. The resulting restriction fragments were resolved by electrophoresis in 2.5% TBE-agarose gels, applying 5,33 V/cm for 1.5 h. The gels were stained with ethidium bromide for visualization under UV light.

    Techniques: Amplification, Sequencing, Agarose Gel Electrophoresis, Polymerase Chain Reaction

    Consensus sequence of the SSUsat monomers from Spisula subtruncata . On the basis of the DNA sequences of the recovered SSUsat monomers from Spisula subtruncata genome, a consensus sequence was derived. Restriction sites for Msp I/ Hpa II, Pvu II and Taq I are underlined. Green and red arrows indicate the positions of PCR primers used for SSUsat amplification in related species.

    Journal: Scientific Reports

    Article Title: Methylation profile of a satellite DNA constituting the intercalary G+C-rich heterochromatin of the cut trough shell Spisula subtruncata (Bivalvia, Mactridae)

    doi: 10.1038/s41598-017-07231-7

    Figure Lengend Snippet: Consensus sequence of the SSUsat monomers from Spisula subtruncata . On the basis of the DNA sequences of the recovered SSUsat monomers from Spisula subtruncata genome, a consensus sequence was derived. Restriction sites for Msp I/ Hpa II, Pvu II and Taq I are underlined. Green and red arrows indicate the positions of PCR primers used for SSUsat amplification in related species.

    Article Snippet: Southern and dot blot hybridisation Genomic DNAs were digested with Hae III (Fermentas), Hpa II (Fermentas), Msp I (Promega), Pvu II (New England Biolabs) and Taq I (Roche).

    Techniques: Sequencing, Derivative Assay, Polymerase Chain Reaction, Amplification

    Representative variation in MSAP profiles. “→” to red arrows represents variation in DNA methylation between diploid and tetraploid plants; “+” represents fragments obtained after digestion with EcoR I or Hpa II/ Msp I; “−” represents fragments not digested by EcoR I or Hpa II/ Msp I; Type I fragments are nonmethylated and were presented in both the H ( EcoR I or Hpa II digest) and M ( EcoR I or Msp I digest) lanes; Type II are fully methylated and only appeared in the M lanes; Type III are hemimethylated and appeared in the H lanes; Type IV were fragments absent from both H and M lanes in diploid but present in either H or M lane of tetraploid, and vice versa.

    Journal: International Journal of Molecular Sciences

    Article Title: Morphological, Genome and Gene Expression Changes in Newly Induced Autopolyploid Chrysanthemum lavandulifolium (Fisch. ex Trautv.) Makino

    doi: 10.3390/ijms17101690

    Figure Lengend Snippet: Representative variation in MSAP profiles. “→” to red arrows represents variation in DNA methylation between diploid and tetraploid plants; “+” represents fragments obtained after digestion with EcoR I or Hpa II/ Msp I; “−” represents fragments not digested by EcoR I or Hpa II/ Msp I; Type I fragments are nonmethylated and were presented in both the H ( EcoR I or Hpa II digest) and M ( EcoR I or Msp I digest) lanes; Type II are fully methylated and only appeared in the M lanes; Type III are hemimethylated and appeared in the H lanes; Type IV were fragments absent from both H and M lanes in diploid but present in either H or M lane of tetraploid, and vice versa.

    Article Snippet: Mthylation Sensitive Amplified Polymorphism Analysis The MSAP technique was applied to the DNA pools from three diploid lines and three tetraploid lines C. lavandulifolium They were digested with either EcoR I and Hpa II or EcoR I and Msp I (NEB) at 37 °C for 12 h. The digested fragments were ligated to 5 pmol EcoR I adaptor and 50 pmol Hpa II/Msp I adaptor by incubation with 4 U T4 DNA polymerase (NEB) at 16 °C for 4 has described for the AFLP method.

    Techniques: DNA Methylation Assay, Methylation

    Comparison of HM-PCR and HM-WGA-PCR results for eleven CGIs . Genomic DNA from peripheral blood leukocytes was digested with Mock (lane 1), Hpa II or Hha I (lane 2), Msp I (lane 3), or McrBC (lane 4). The digested genomic DNA were used for PCR amplification either directly (left; HM-PCR) or after whole-genome-amplification (right; HM-WGA-PCR) using primer pairs for the eight CGIs on chromosome 21 (A~D) and three CGIs on chromosome 11 (E). Here, when Hha I-digested genomic DNA was used in lane 2, 1 ul of distilled water was used in lane 3 in place of Msp I-digested genomic DNA. PCR products were electrophoresed, stained with ethidium bromide, and visualized by UV illumination. Results of bisulfite sequencing are shown for the eight CGIs on chromosome 21 (A~D). Open and closed circles indicate unmethylated and methylated CpG dinucleotides, respectively. Each row of circles represents each sequenced clone of bisufite PCR products.

    Journal: BMC Research Notes

    Article Title: Highly efficient PCR assay to discriminate allelic DNA methylation status using whole genome amplification

    doi: 10.1186/1756-0500-4-179

    Figure Lengend Snippet: Comparison of HM-PCR and HM-WGA-PCR results for eleven CGIs . Genomic DNA from peripheral blood leukocytes was digested with Mock (lane 1), Hpa II or Hha I (lane 2), Msp I (lane 3), or McrBC (lane 4). The digested genomic DNA were used for PCR amplification either directly (left; HM-PCR) or after whole-genome-amplification (right; HM-WGA-PCR) using primer pairs for the eight CGIs on chromosome 21 (A~D) and three CGIs on chromosome 11 (E). Here, when Hha I-digested genomic DNA was used in lane 2, 1 ul of distilled water was used in lane 3 in place of Msp I-digested genomic DNA. PCR products were electrophoresed, stained with ethidium bromide, and visualized by UV illumination. Results of bisulfite sequencing are shown for the eight CGIs on chromosome 21 (A~D). Open and closed circles indicate unmethylated and methylated CpG dinucleotides, respectively. Each row of circles represents each sequenced clone of bisufite PCR products.

    Article Snippet: HM-WGA-PCR Genomic DNA (500 ng) was digested with 50 U of Hpa II, Hha I, Msp I (TaKaRa), or McrBC (New England Biolabs) overnight at 37°C in 50 μl of the recommended buffers.

    Techniques: Polymerase Chain Reaction, Whole Genome Amplification, Amplification, Staining, Methylation Sequencing, Methylation

    Principles of the Hpa II- McrBC whole-genome-amplification PCR (HM-WGA-PCR) method . The two parallel lines in the

    Journal: BMC Research Notes

    Article Title: Highly efficient PCR assay to discriminate allelic DNA methylation status using whole genome amplification

    doi: 10.1186/1756-0500-4-179

    Figure Lengend Snippet: Principles of the Hpa II- McrBC whole-genome-amplification PCR (HM-WGA-PCR) method . The two parallel lines in the "Digestion" panel indicate genomic amplicons from both alleles. The circled "m" indicates a methylated CpG dinucleotide. Hpa II digests unmethylated CCGG, but not methylated C m CGG. In contrast, McrBC digests methylated R m C 40-80 R m C sequences, but not unmethylated RC 40-80 RC. Genomic DNA is digested with Hpa II and McrBC independently. Subsequently, an aliquot of each restriction-enzyme-digested DNA (50 ng) is subjected to whole-genome-amplification (WGA) to yield 5 μg of whole-genome-amplified DNA. Using an aliquot of the amplified DNA (50 ng), the target DNA region is PCR-amplified by the primer pair (dotted arrows). The PCR products from the Hpa II/ Hha I-digested and McrBC -digested DNA are electrophoresed, stained with ethidium bromide, and visualized by UV illumination. If an amplicon is fully methylated (i.e., complete methylation), it is digested by McrBC , but not by Hpa II. Thus, it is amplified from the Hpa II-digested and whole-genome-amplified DNA, but not from the McrBC -digested and whole-genome-amplified DNA. By contrast, if an amplicon totally escapes methylation (i.e., null methylation), it is digested by Hpa II, but not by McrBC . Thus, it is amplified from McrBC -digested and whole-genome-amplified DNA, but not from Hpa II-digested and whole-genome-amplified DNA. If an amplicon contains both methylated and unmethylated alleles (i.e., composite methylation), it is amplified from both whole-genome-amplified templates. If an amplicon is partially methylated on both alleles (i.e., incomplete methylation), it is amplified from neither whole-genome-amplified template.

    Article Snippet: HM-WGA-PCR Genomic DNA (500 ng) was digested with 50 U of Hpa II, Hha I, Msp I (TaKaRa), or McrBC (New England Biolabs) overnight at 37°C in 50 μl of the recommended buffers.

    Techniques: Whole Genome Amplification, Polymerase Chain Reaction, Methylation, Amplification, Staining

    Effect of combined DDM1 and siRNA deficiency on 5S rDNA methylation and aberrant transcripts. A) 5S rDNA hypermethylation in ddm1 is siRNA-dependent: Southern blot comparison of Alu I and Hae III-digested genomic DNA isolated from inflorescences of wild type (WT), ddm1 , and double mutant lines nrpd1 ddm1 , rdr2 ddm1 , and dcl3 ddm1 (top panel). The probe is the same as in Figure 2E . Dilutions of the above digests were also assayed by PCR using 5S LT1 primers (bottom panel). Samples to which no restriction enzyme was added are controls (no digest). B) 5S LT1 is silenced by two overlapping processes: RNA samples from inflorescences of WT, ddm1 and the double mutant panel were analyzed by one-step RT-PCR, performed as described in Figure 1C . Control reactions were performed with ACT2 primers; reverse transcriptase was omitted from duplicate 5S LT1 and ACT2 reactions (no RT).

    Journal: PLoS ONE

    Article Title: Heterochromatic siRNAs and DDM1 Independently Silence Aberrant 5S rDNA Transcripts in Arabidopsis

    doi: 10.1371/journal.pone.0005932

    Figure Lengend Snippet: Effect of combined DDM1 and siRNA deficiency on 5S rDNA methylation and aberrant transcripts. A) 5S rDNA hypermethylation in ddm1 is siRNA-dependent: Southern blot comparison of Alu I and Hae III-digested genomic DNA isolated from inflorescences of wild type (WT), ddm1 , and double mutant lines nrpd1 ddm1 , rdr2 ddm1 , and dcl3 ddm1 (top panel). The probe is the same as in Figure 2E . Dilutions of the above digests were also assayed by PCR using 5S LT1 primers (bottom panel). Samples to which no restriction enzyme was added are controls (no digest). B) 5S LT1 is silenced by two overlapping processes: RNA samples from inflorescences of WT, ddm1 and the double mutant panel were analyzed by one-step RT-PCR, performed as described in Figure 1C . Control reactions were performed with ACT2 primers; reverse transcriptase was omitted from duplicate 5S LT1 and ACT2 reactions (no RT).

    Article Snippet: 3 µg aliquots of DNA were digested overnight with 30 U of Alu I, Hpa II or Hae III in the corresponding commercial buffer (New England Biolabs).

    Techniques: Methylation, Southern Blot, Isolation, Mutagenesis, Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction

    DDM1 limits IGS siRNA accumulation and asymmetric methylation. A) Detection of IGS siRNAs (siR1003) in dicer-like ( dcl ) mutant combinations. Blot analysis of small RNA isolated from leaves of wild type (WT); single mutants dcl2, dcl3, dcl4 ; double mutants dcl2 dcl3 ( dcl2/3 ), dcl2 dcl4 ( dcl2/4 ), dcl3 dcl4 ( dcl3/4 ); and the triple mutant dcl2 dcl3 dcl4 ( dcl2/3/4 ). B) Localization of IGS-related RNA in interphase nuclei by fluorescent in situ hybridization with an RNA probe for siR1003 (red). DNA was stained with DAPI (white). The size bar corresponds to 5 µm. C) Overaccumulation of IGS siRNAs in the ddm1 background. Blot analysis of small RNA from dcl3 and double mutant dcl3 ddm1 using probes for siR1003 and the larger 3′flanking region ( Figure 1A , diagram). WT and ddm1 signals from the same membrane are provided for comparison. D) IGS siRNA overaccumulation persists in out-crossed ddm1 . Blot analysis of small RNA isolated from WT, nrpd1 (−/−), ddm1 (−/−) and F1 heterozygotes (+/−) of each mutant crossed to the WT. E) Asymmetric cytosines in the IGS are hypermethylated in met1 and ddm1 . Southern blot analysis was performed on Alu I-digested genomic DNA isolated from WT, nrpd1 , rdr2 , two alleles of dcl3 , met1 and ddm1 . The probe corresponds to 5S LT1, shown aligned to a representative 5S rDNA repeat unit from Chromosome 5. Alu I and Hae III sites in the IGS region are indicated.

    Journal: PLoS ONE

    Article Title: Heterochromatic siRNAs and DDM1 Independently Silence Aberrant 5S rDNA Transcripts in Arabidopsis

    doi: 10.1371/journal.pone.0005932

    Figure Lengend Snippet: DDM1 limits IGS siRNA accumulation and asymmetric methylation. A) Detection of IGS siRNAs (siR1003) in dicer-like ( dcl ) mutant combinations. Blot analysis of small RNA isolated from leaves of wild type (WT); single mutants dcl2, dcl3, dcl4 ; double mutants dcl2 dcl3 ( dcl2/3 ), dcl2 dcl4 ( dcl2/4 ), dcl3 dcl4 ( dcl3/4 ); and the triple mutant dcl2 dcl3 dcl4 ( dcl2/3/4 ). B) Localization of IGS-related RNA in interphase nuclei by fluorescent in situ hybridization with an RNA probe for siR1003 (red). DNA was stained with DAPI (white). The size bar corresponds to 5 µm. C) Overaccumulation of IGS siRNAs in the ddm1 background. Blot analysis of small RNA from dcl3 and double mutant dcl3 ddm1 using probes for siR1003 and the larger 3′flanking region ( Figure 1A , diagram). WT and ddm1 signals from the same membrane are provided for comparison. D) IGS siRNA overaccumulation persists in out-crossed ddm1 . Blot analysis of small RNA isolated from WT, nrpd1 (−/−), ddm1 (−/−) and F1 heterozygotes (+/−) of each mutant crossed to the WT. E) Asymmetric cytosines in the IGS are hypermethylated in met1 and ddm1 . Southern blot analysis was performed on Alu I-digested genomic DNA isolated from WT, nrpd1 , rdr2 , two alleles of dcl3 , met1 and ddm1 . The probe corresponds to 5S LT1, shown aligned to a representative 5S rDNA repeat unit from Chromosome 5. Alu I and Hae III sites in the IGS region are indicated.

    Article Snippet: 3 µg aliquots of DNA were digested overnight with 30 U of Alu I, Hpa II or Hae III in the corresponding commercial buffer (New England Biolabs).

    Techniques: Methylation, Mutagenesis, Isolation, In Situ Hybridization, Staining, Southern Blot