hpy188i Search Results


  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 96
    New England Biolabs hpy 188i
    c idA _IV gene polymorphic regions and PCR-RFLP tests for specific variants. a Schematic representation of the architecture of cidA _IV polymorphism. The black line represents the cidA_IV sequence with a focus on the two polymorphic regions. Non-polymorphic regions were shortened and represented as a dashed line. Numbers in red under the line represent nucleotide positions, and the numbers in blue indicate amino-acid positions. The overlapping oligonucleotides used for PCR amplification are represented by arrows numbered as in Supplementary Table 2 . Both polymorphic regions were studied, but only the upstream matched the compatibility profile; its four different sequences (α, β γ, and δ) were followed by one of the two possible sequences (1 or 2) in the downstream polymorphic region. A different color code was used for α (purple), β (light blue), γ (light green), and δ (yellow) sequences in the upstream part of the cidA gene. b The repertoire of cidA _IV variants is different in “compatible” and “incompatible” lines. cidA _IV(δ) is present only in lines with “incompatible” crossing type. The names of the C . pipiens lines used to set-up the PCR-RFLP ( c ) test are highlighted in red. c PCR-RFLP tests for distinguishing between cidA _IV variants on the basis of the upstream polymorphic region. A 778 bp fragment was amplified with primers 1/13. Double digestion with Apo I and <t>Hpy</t> <t>188I</t> distinguished between cidA _IV(α) (six fragments: 471; 122; 57; 53; 51; and 24 bp), cidA _IV(β) and cidA _IV(γ) (six fragments: 441; 122; 83; 57; 51; and 24 bp), and cidA _IV(δ) (five fragments: 524; 122; 57; 51; and 24 bp). Panel to the left of the electrophoresis gel: PCR-RFLP on DNA from clones; right panel: PCR-RFLP on DNA from the Istanbul, Harash, Ichkeul 13, and Ichkeul 09 lines. On the right of the gel, a schematic representation of the PCR-RFLP profiles of these lines. Digestion bands that are specific of the variants are represented with the color code established in a . Bands that are not used to discriminate between the variants are represented in black on the bottom of the schematic gel
    Hpy 188i, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 96/100, based on 98 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hpy 188i/product/New England Biolabs
    Average 96 stars, based on 98 article reviews
    Price from $9.99 to $1999.99
    hpy 188i - by Bioz Stars, 2020-08
    96/100 stars
      Buy from Supplier

    90
    Thermo Fisher hpy 188i restriction enzyme
    IFITM3 rs3888188 T > G polymorphism using polymerase chain reaction-restriction fragment length polymorphism methods. The T allele was undigested (586-bp), while the G allele was digested by the <t>Hpy</t> <t>188I</t> restriction enzyme and produces 340- and 246-bp
    Hpy 188i Restriction Enzyme, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hpy 188i restriction enzyme/product/Thermo Fisher
    Average 90 stars, based on 3 article reviews
    Price from $9.99 to $1999.99
    hpy 188i restriction enzyme - by Bioz Stars, 2020-08
    90/100 stars
      Buy from Supplier

    91
    Thermo Fisher hpy 188i
    IFITM3 rs3888188 T > G polymorphism using polymerase chain reaction-restriction fragment length polymorphism methods. The T allele was undigested (586-bp), while the G allele was digested by the <t>Hpy</t> <t>188I</t> restriction enzyme and produces 340- and 246-bp
    Hpy 188i, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 91/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hpy 188i/product/Thermo Fisher
    Average 91 stars, based on 4 article reviews
    Price from $9.99 to $1999.99
    hpy 188i - by Bioz Stars, 2020-08
    91/100 stars
      Buy from Supplier

    99
    New England Biolabs restriction endonuclease hpy188i
    IFITM3 rs3888188 T > G polymorphism using polymerase chain reaction-restriction fragment length polymorphism methods. The T allele was undigested (586-bp), while the G allele was digested by the <t>Hpy</t> <t>188I</t> restriction enzyme and produces 340- and 246-bp
    Restriction Endonuclease Hpy188i, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/restriction endonuclease hpy188i/product/New England Biolabs
    Average 99 stars, based on 10 article reviews
    Price from $9.99 to $1999.99
    restriction endonuclease hpy188i - by Bioz Stars, 2020-08
    99/100 stars
      Buy from Supplier

    Image Search Results


    c idA _IV gene polymorphic regions and PCR-RFLP tests for specific variants. a Schematic representation of the architecture of cidA _IV polymorphism. The black line represents the cidA_IV sequence with a focus on the two polymorphic regions. Non-polymorphic regions were shortened and represented as a dashed line. Numbers in red under the line represent nucleotide positions, and the numbers in blue indicate amino-acid positions. The overlapping oligonucleotides used for PCR amplification are represented by arrows numbered as in Supplementary Table 2 . Both polymorphic regions were studied, but only the upstream matched the compatibility profile; its four different sequences (α, β γ, and δ) were followed by one of the two possible sequences (1 or 2) in the downstream polymorphic region. A different color code was used for α (purple), β (light blue), γ (light green), and δ (yellow) sequences in the upstream part of the cidA gene. b The repertoire of cidA _IV variants is different in “compatible” and “incompatible” lines. cidA _IV(δ) is present only in lines with “incompatible” crossing type. The names of the C . pipiens lines used to set-up the PCR-RFLP ( c ) test are highlighted in red. c PCR-RFLP tests for distinguishing between cidA _IV variants on the basis of the upstream polymorphic region. A 778 bp fragment was amplified with primers 1/13. Double digestion with Apo I and Hpy 188I distinguished between cidA _IV(α) (six fragments: 471; 122; 57; 53; 51; and 24 bp), cidA _IV(β) and cidA _IV(γ) (six fragments: 441; 122; 83; 57; 51; and 24 bp), and cidA _IV(δ) (five fragments: 524; 122; 57; 51; and 24 bp). Panel to the left of the electrophoresis gel: PCR-RFLP on DNA from clones; right panel: PCR-RFLP on DNA from the Istanbul, Harash, Ichkeul 13, and Ichkeul 09 lines. On the right of the gel, a schematic representation of the PCR-RFLP profiles of these lines. Digestion bands that are specific of the variants are represented with the color code established in a . Bands that are not used to discriminate between the variants are represented in black on the bottom of the schematic gel

    Journal: Nature Communications

    Article Title: Culex pipiens crossing type diversity is governed by an amplified and polymorphic operon of Wolbachia

    doi: 10.1038/s41467-017-02749-w

    Figure Lengend Snippet: c idA _IV gene polymorphic regions and PCR-RFLP tests for specific variants. a Schematic representation of the architecture of cidA _IV polymorphism. The black line represents the cidA_IV sequence with a focus on the two polymorphic regions. Non-polymorphic regions were shortened and represented as a dashed line. Numbers in red under the line represent nucleotide positions, and the numbers in blue indicate amino-acid positions. The overlapping oligonucleotides used for PCR amplification are represented by arrows numbered as in Supplementary Table 2 . Both polymorphic regions were studied, but only the upstream matched the compatibility profile; its four different sequences (α, β γ, and δ) were followed by one of the two possible sequences (1 or 2) in the downstream polymorphic region. A different color code was used for α (purple), β (light blue), γ (light green), and δ (yellow) sequences in the upstream part of the cidA gene. b The repertoire of cidA _IV variants is different in “compatible” and “incompatible” lines. cidA _IV(δ) is present only in lines with “incompatible” crossing type. The names of the C . pipiens lines used to set-up the PCR-RFLP ( c ) test are highlighted in red. c PCR-RFLP tests for distinguishing between cidA _IV variants on the basis of the upstream polymorphic region. A 778 bp fragment was amplified with primers 1/13. Double digestion with Apo I and Hpy 188I distinguished between cidA _IV(α) (six fragments: 471; 122; 57; 53; 51; and 24 bp), cidA _IV(β) and cidA _IV(γ) (six fragments: 441; 122; 83; 57; 51; and 24 bp), and cidA _IV(δ) (five fragments: 524; 122; 57; 51; and 24 bp). Panel to the left of the electrophoresis gel: PCR-RFLP on DNA from clones; right panel: PCR-RFLP on DNA from the Istanbul, Harash, Ichkeul 13, and Ichkeul 09 lines. On the right of the gel, a schematic representation of the PCR-RFLP profiles of these lines. Digestion bands that are specific of the variants are represented with the color code established in a . Bands that are not used to discriminate between the variants are represented in black on the bottom of the schematic gel

    Article Snippet: Double digestion of the PCR products with Apo I and Hpy 188I (New England Biolabs) identified three groups of sequencing: cid A_IV(α) (six fragments: 471; 122; 57; 53; 51; and 24 bp); cidA _IV(β) and cidA _IV(γ) (six fragments: 441; 123; 83; 57; 51; and 24 bp); and cidA _IV(δ) (five fragments: 524; 122; 57; 51; and 24 bp).

    Techniques: Polymerase Chain Reaction, Sequencing, Amplification, Electrophoresis, Clone Assay

    CRISPR/Cas9-mediated generation of Dmc1 M200V mice. ( A ) Diagram of human DMC1 protein labeled with known functional domains and binding sites. The M200V amino acid change is encoded by SNP rs2227914. ( B ) CRISPR-Cas9 genome editing strategy to introduce the M200V amino acid change. Underlined is the sgRNA sequence, in bold is the PAM site, highlighted in yellow are nucleotide changes for Val and italicized is the restriction enzyme site for Hpy 188I (TCN/GA). ( C ) Sanger sequencing chromatograms from WT (left) and Dmc1 M200V/M200V mouse (right).

    Journal: Human Molecular Genetics

    Article Title: A putative human infertility allele of the meiotic recombinase DMC1 does not affect fertility in mice

    doi: 10.1093/hmg/ddy286

    Figure Lengend Snippet: CRISPR/Cas9-mediated generation of Dmc1 M200V mice. ( A ) Diagram of human DMC1 protein labeled with known functional domains and binding sites. The M200V amino acid change is encoded by SNP rs2227914. ( B ) CRISPR-Cas9 genome editing strategy to introduce the M200V amino acid change. Underlined is the sgRNA sequence, in bold is the PAM site, highlighted in yellow are nucleotide changes for Val and italicized is the restriction enzyme site for Hpy 188I (TCN/GA). ( C ) Sanger sequencing chromatograms from WT (left) and Dmc1 M200V/M200V mouse (right).

    Article Snippet: For identification of Dmc1M200V mice, PCR samples were digested with restriction enzyme Hpy 188I (NEB) at 37°C for 2 h then analyzed using agarose gel electrophoresis.

    Techniques: CRISPR, Mouse Assay, Labeling, Functional Assay, Binding Assay, Introduce, Sequencing

    Testing for the 4qA161 haplotype by differential digestion with Hpy188I and Hpy188III. Genomic DNAs and human-rodent somatic cell hybrids (GM14193, GM11687, and GM10926 containing chr4, chr4, and chr10, respectively; Coriell Institute) were analysed for overlapping Hpy188I (purple) or Hpy188III (blue) sites. Whether one or two 4qA161 alleles were present was subsequently determined by the sequencing assay. GM11448 and GM10115 containing chr4 were negative for 4qA161 (not shown) (A). The Hpy188I or Hpy188III sites are indicated for the forward strand in the SNP-rich region of 4qA161, the other main 4qA alleles, and the predominant 10qA allele (B).

    Journal: Journal of medical genetics

    Article Title: FSH dystrophy and a subtelomeric 4q haplotype: a new assay and associations with disease

    doi: 10.1136/jmg.2009.076703

    Figure Lengend Snippet: Testing for the 4qA161 haplotype by differential digestion with Hpy188I and Hpy188III. Genomic DNAs and human-rodent somatic cell hybrids (GM14193, GM11687, and GM10926 containing chr4, chr4, and chr10, respectively; Coriell Institute) were analysed for overlapping Hpy188I (purple) or Hpy188III (blue) sites. Whether one or two 4qA161 alleles were present was subsequently determined by the sequencing assay. GM11448 and GM10115 containing chr4 were negative for 4qA161 (not shown) (A). The Hpy188I or Hpy188III sites are indicated for the forward strand in the SNP-rich region of 4qA161, the other main 4qA alleles, and the predominant 10qA allele (B).

    Article Snippet: Aliquots of the single 168-bp PCR product (10 µl) were digested with 20 U of Hpy188I or of Hpy188III (New England Biolabs) for 3 h at 37°C (40 µl reaction volume).

    Techniques: Sequencing

    Mutation analysis of GJA8 . Sequence chromatograms of the wild-type allele ( A ) demonstrate a nucleotide sequence encoding a serine (Ser) at codon 276, and the chromatograms of the mutant allele ( B ) demonstrate a C to T transition resulting in the substitution of serine (Ser) by phenylalanine (Phe). Confirmation of segregation of the S276F mutation was given by the PCR-RFLP method ( C ). M is the DNA ladder; Lane 1 shows the digestions of the PCR products amplified from samples of the patients by Hpy188I. Lane 2 and 4 show the PCR products amplified from samples of one patient or the unaffected member in the pedigree. Lane 3 illustrates the digestions of the PCR products amplified from samples of the unaffected family member by Hpy188I. The mutant primer results in the gain of Hpy 188I sites producing digested fragments of 215 and 24 bp with wild-type GJA8 alleles, and the mutation in the GJA8 gene leads to an abolition of this site, remaining undigested (239 bp).

    Journal: Molecular Vision

    Article Title: A novel connexin 50 (GJA8) mutation in a Chinese family with a dominant congenital pulverulent nuclear cataract

    doi:

    Figure Lengend Snippet: Mutation analysis of GJA8 . Sequence chromatograms of the wild-type allele ( A ) demonstrate a nucleotide sequence encoding a serine (Ser) at codon 276, and the chromatograms of the mutant allele ( B ) demonstrate a C to T transition resulting in the substitution of serine (Ser) by phenylalanine (Phe). Confirmation of segregation of the S276F mutation was given by the PCR-RFLP method ( C ). M is the DNA ladder; Lane 1 shows the digestions of the PCR products amplified from samples of the patients by Hpy188I. Lane 2 and 4 show the PCR products amplified from samples of one patient or the unaffected member in the pedigree. Lane 3 illustrates the digestions of the PCR products amplified from samples of the unaffected family member by Hpy188I. The mutant primer results in the gain of Hpy 188I sites producing digested fragments of 215 and 24 bp with wild-type GJA8 alleles, and the mutation in the GJA8 gene leads to an abolition of this site, remaining undigested (239 bp).

    Article Snippet: If there was a mutation of C to T, the recognition site of restriction enzyme Hpy188I would have been abolished.

    Techniques: Mutagenesis, Sequencing, Polymerase Chain Reaction, Amplification

    Genomic disruption of the β-catenin gene ( CTNNB1 ) in BxPC-3 cells using ZFNs. A) Outline of the CTNNB1 gene with the N-terminal (NTD) and C-terminal (CTD) domains colored in brown and blue, respectively. Binding sequence for ZFN-1 and -2 (green) flanking the spacer region (red) incorporating the recognition site for the Hpy188I restriction endonuclease are shown with the corresponding codon number in the CTNNB1 gene below. B) β-catenin immunostaining of BxPC-3 wild type (WT) and CTNNB1 gene disrupted clones (#4, #31, #79, #93 and #111) with DAPI nuclear counterstaining. In the wild type cells β-catenin is predominantly localized at the cell membranes. C) Western blot analysis of WT and gene disrupted clones using β-catenin (CTD) and actin specific Abs. For wild type cells 5 µg of total protein (TP) and for the gene disrupted clones 20 µg of TP was applied for each lane. The position and molecular weight of the β-catenin and actin bands are indicated.

    Journal: PLoS ONE

    Article Title: Implications of Targeted Genomic Disruption of β-Catenin in BxPC-3 Pancreatic Adenocarcinoma Cells

    doi: 10.1371/journal.pone.0115496

    Figure Lengend Snippet: Genomic disruption of the β-catenin gene ( CTNNB1 ) in BxPC-3 cells using ZFNs. A) Outline of the CTNNB1 gene with the N-terminal (NTD) and C-terminal (CTD) domains colored in brown and blue, respectively. Binding sequence for ZFN-1 and -2 (green) flanking the spacer region (red) incorporating the recognition site for the Hpy188I restriction endonuclease are shown with the corresponding codon number in the CTNNB1 gene below. B) β-catenin immunostaining of BxPC-3 wild type (WT) and CTNNB1 gene disrupted clones (#4, #31, #79, #93 and #111) with DAPI nuclear counterstaining. In the wild type cells β-catenin is predominantly localized at the cell membranes. C) Western blot analysis of WT and gene disrupted clones using β-catenin (CTD) and actin specific Abs. For wild type cells 5 µg of total protein (TP) and for the gene disrupted clones 20 µg of TP was applied for each lane. The position and molecular weight of the β-catenin and actin bands are indicated.

    Article Snippet: For RFLP analysis the PCR products from CTNNB1 exon 3 were purified (illustra GFX PCR DNA and Gel Band Purification Kit, GE Healthcare) and digested with Hpy188I restriction enzyme (New England Biolabs) before separation on 4% agarose gel (Agarose 1000, Invitrogen, CA, USA).

    Techniques: Binding Assay, Sequencing, Immunostaining, Clone Assay, Western Blot, Molecular Weight

    Characterization of genomic changes present in the CTNNB1 gene disrupted cells. A) PCR analysis of the integrity of exon 3 in the CTNNB1 gene using genomic DNA isolated from WT and gene disrupted clones (#4, #31, #79, #93 and #111) as template. ctr: negative control devoid of gDNA, M: molecular size marker. B) Genotyping of WT and gene disrupted clones by RFLP. Digestion of PCR products spanning exon 3 of the CTNNB1 gene with the Hpy188I restriction endonuclease that would cut undisrupted wild type sequences. u: PCR product without Hpy188I, c: PCR product with Hpy188I, M: molecular size marker. C) Sequencing of PCR products spanning exon 3 of CTNNB1 gene from WT and gene disrupted clones. The binding sequence for ZFN-1 and -2 (green) flanking the spacer region (red) is indicated above. * in the sequence indicates deleted base and the total number of deleted bases are shown at the right. D) PCR analysis of exon integrity in WT and gene disrupted clones #4 and #111 using primers designed for amplifying the promoter region (Prom) and exons 2, 3, 4, 5, 7 and 12 (Ex-2, 3, 4, 5, 7 and 12 respectively). Expected sizes of amplification products of intact sequences are indicated. M: molecular size marker. Upper panel: Schematic representation of the CTNNB1 gene structure.

    Journal: PLoS ONE

    Article Title: Implications of Targeted Genomic Disruption of β-Catenin in BxPC-3 Pancreatic Adenocarcinoma Cells

    doi: 10.1371/journal.pone.0115496

    Figure Lengend Snippet: Characterization of genomic changes present in the CTNNB1 gene disrupted cells. A) PCR analysis of the integrity of exon 3 in the CTNNB1 gene using genomic DNA isolated from WT and gene disrupted clones (#4, #31, #79, #93 and #111) as template. ctr: negative control devoid of gDNA, M: molecular size marker. B) Genotyping of WT and gene disrupted clones by RFLP. Digestion of PCR products spanning exon 3 of the CTNNB1 gene with the Hpy188I restriction endonuclease that would cut undisrupted wild type sequences. u: PCR product without Hpy188I, c: PCR product with Hpy188I, M: molecular size marker. C) Sequencing of PCR products spanning exon 3 of CTNNB1 gene from WT and gene disrupted clones. The binding sequence for ZFN-1 and -2 (green) flanking the spacer region (red) is indicated above. * in the sequence indicates deleted base and the total number of deleted bases are shown at the right. D) PCR analysis of exon integrity in WT and gene disrupted clones #4 and #111 using primers designed for amplifying the promoter region (Prom) and exons 2, 3, 4, 5, 7 and 12 (Ex-2, 3, 4, 5, 7 and 12 respectively). Expected sizes of amplification products of intact sequences are indicated. M: molecular size marker. Upper panel: Schematic representation of the CTNNB1 gene structure.

    Article Snippet: For RFLP analysis the PCR products from CTNNB1 exon 3 were purified (illustra GFX PCR DNA and Gel Band Purification Kit, GE Healthcare) and digested with Hpy188I restriction enzyme (New England Biolabs) before separation on 4% agarose gel (Agarose 1000, Invitrogen, CA, USA).

    Techniques: Polymerase Chain Reaction, Isolation, Clone Assay, Negative Control, Marker, Sequencing, Binding Assay, Amplification

    REA patterns obtained after Hpy188-III digestion of the PCR product of the gyrB - 2 fragment (897 bp). Lanes: 1 and 6, 100-bp-ladder molecular size markers; 2 and 3, digested PCR products from type III strains; 4 and 5, digested PCR products from type I and II strains, respectively.

    Journal: Journal of Clinical Microbiology

    Article Title: Polymorphisms in gyrA and gyrB Genes among Mycobacterium avium subsp. paratuberculosis Type I, II, and III Isolates ▿

    doi: 10.1128/JCM.01411-07

    Figure Lengend Snippet: REA patterns obtained after Hpy188-III digestion of the PCR product of the gyrB - 2 fragment (897 bp). Lanes: 1 and 6, 100-bp-ladder molecular size markers; 2 and 3, digested PCR products from type III strains; 4 and 5, digested PCR products from type I and II strains, respectively.

    Article Snippet: The 897-bp PCR products were submitted to restriction analysis with the enzyme Hpy188-III (New England Biolabs).

    Techniques: Polymerase Chain Reaction

    Restriction enzyme (RE)-assisted methylation analysis of the proximal MOR promoter from dissected mouse brain regions. Analyses were performed as described for Fig. 3 , except as indicated. (A) – For the –569 CpG site, methylation-specific PCR products were amplified using MS-630 and MAS-180 primers (Table 1 ), generating the Full-A fragment. Hp1 was used to cut the amplified PCR products. For the –182 and –178/–170 sites, PCR amplification was performed with the MS-376 and MAS-79 primers (Full-B) and samples were digested with either Hp1 or Hp4, respectively. (B) – Methylation-specific PCR products from various regions of the mouse brain. The demethylation of the MOR promoter is expressed graphically below the gel as the mean ± S.E.M. (C) and (D) – RE-assisted methylation analysis of the MOR DP from dissected mouse brain regions. Methylation-specific PCR products were amplified using MS-1754 and MAS-927 primers (Table 1 ) generating the Full fragment. Several Restriction enzymes (ClaI, BstBI, and HpyCH4IV) were used to detect methylation status, as indicated. The analyses and quantification of methylation were performed as described above. (E)– Methylation status of the DP of the MOR gene in mouse brain regions (olfactory bulb, striatum, hypothalamus, and cerebellum). Except as noted below, experiments were performed as described for Fig. 1D . dpTIS: transcription initiation site of the MOR DP. Methylation-specific PCR was performed with primers MS-1754 and MAS-927 (see Table 1 ) followed by TA cloning (Invitrogen). The percentages of methyl CpG versus unmethylated CpG are indicated for all CpG sites.

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: Epigenetic programming of μ-opioid receptor gene in mouse brain is regulated by MeCP2 and brg1 chromatin remodelling factor

    doi: 10.1111/j.1582-4934.2008.00535.x

    Figure Lengend Snippet: Restriction enzyme (RE)-assisted methylation analysis of the proximal MOR promoter from dissected mouse brain regions. Analyses were performed as described for Fig. 3 , except as indicated. (A) – For the –569 CpG site, methylation-specific PCR products were amplified using MS-630 and MAS-180 primers (Table 1 ), generating the Full-A fragment. Hp1 was used to cut the amplified PCR products. For the –182 and –178/–170 sites, PCR amplification was performed with the MS-376 and MAS-79 primers (Full-B) and samples were digested with either Hp1 or Hp4, respectively. (B) – Methylation-specific PCR products from various regions of the mouse brain. The demethylation of the MOR promoter is expressed graphically below the gel as the mean ± S.E.M. (C) and (D) – RE-assisted methylation analysis of the MOR DP from dissected mouse brain regions. Methylation-specific PCR products were amplified using MS-1754 and MAS-927 primers (Table 1 ) generating the Full fragment. Several Restriction enzymes (ClaI, BstBI, and HpyCH4IV) were used to detect methylation status, as indicated. The analyses and quantification of methylation were performed as described above. (E)– Methylation status of the DP of the MOR gene in mouse brain regions (olfactory bulb, striatum, hypothalamus, and cerebellum). Except as noted below, experiments were performed as described for Fig. 1D . dpTIS: transcription initiation site of the MOR DP. Methylation-specific PCR was performed with primers MS-1754 and MAS-927 (see Table 1 ) followed by TA cloning (Invitrogen). The percentages of methyl CpG versus unmethylated CpG are indicated for all CpG sites.

    Article Snippet: All the methyl-CpG-sensitive restriction enzymes (BstBI, ClaI, Hpy188I and HpyCH4IV) were purchased from New England Biolabs (Ipswich, MA, USA).

    Techniques: Methylation, Polymerase Chain Reaction, Amplification, Mass Spectrometry, TA Cloning

    Analysis of DNA methylation by methylation-sensitive restriction enzyme assay. (A) – The proximal MOR promoter, its flanking regions, and the positions of the CpG sites are shown. Methylation-specific PCR products (Full-A) were amplified using MS-647 and MAS-353 primers (Table 1 ) after sodium bisulphite treatment for genomic DNA. The restriction enzyme Hpy188I (Hp1) recognizes and cuts amplified PCR products that contain the retained C residue at the –569 position. For the –182 and –178/–170 sites, MS-376 and MAS-79 primers were used for the PCR amplification (Full-B) and digested with either Hp1 or HpyCH4IV (Hp4), respectively. The predicted sizes of the methylation-specific PCR products and the digested fragments are shown below. (B) – PCR products (Full-A and Full-B fragments) from NS20Y cells treated with the indicated concentrations of 5-aza-dC, as well as from undifferentiated P19 cells, fully differentiated P19 cells (AP4d), and mouse brain (MB). Intact PCR product (no RE) was used as a control. The graphs to the right of each gel show the percentage of methylation, determined by comparing the intensity of the bands cut by Hp1 or Hp4 relative to the intensity of the unmethylated ( i.e. upper, uncut bands). Column numbers in graph indicate the corresponding gel lanes. Data for the graphs are the mean ± S.E.M. from three independent experiments. *Indicates P

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: Epigenetic programming of μ-opioid receptor gene in mouse brain is regulated by MeCP2 and brg1 chromatin remodelling factor

    doi: 10.1111/j.1582-4934.2008.00535.x

    Figure Lengend Snippet: Analysis of DNA methylation by methylation-sensitive restriction enzyme assay. (A) – The proximal MOR promoter, its flanking regions, and the positions of the CpG sites are shown. Methylation-specific PCR products (Full-A) were amplified using MS-647 and MAS-353 primers (Table 1 ) after sodium bisulphite treatment for genomic DNA. The restriction enzyme Hpy188I (Hp1) recognizes and cuts amplified PCR products that contain the retained C residue at the –569 position. For the –182 and –178/–170 sites, MS-376 and MAS-79 primers were used for the PCR amplification (Full-B) and digested with either Hp1 or HpyCH4IV (Hp4), respectively. The predicted sizes of the methylation-specific PCR products and the digested fragments are shown below. (B) – PCR products (Full-A and Full-B fragments) from NS20Y cells treated with the indicated concentrations of 5-aza-dC, as well as from undifferentiated P19 cells, fully differentiated P19 cells (AP4d), and mouse brain (MB). Intact PCR product (no RE) was used as a control. The graphs to the right of each gel show the percentage of methylation, determined by comparing the intensity of the bands cut by Hp1 or Hp4 relative to the intensity of the unmethylated ( i.e. upper, uncut bands). Column numbers in graph indicate the corresponding gel lanes. Data for the graphs are the mean ± S.E.M. from three independent experiments. *Indicates P

    Article Snippet: All the methyl-CpG-sensitive restriction enzymes (BstBI, ClaI, Hpy188I and HpyCH4IV) were purchased from New England Biolabs (Ipswich, MA, USA).

    Techniques: DNA Methylation Assay, Methylation, Enzymatic Assay, Polymerase Chain Reaction, Amplification, Mass Spectrometry

    IFITM3 rs3888188 T > G polymorphism using polymerase chain reaction-restriction fragment length polymorphism methods. The T allele was undigested (586-bp), while the G allele was digested by the Hpy 188I restriction enzyme and produces 340- and 246-bp

    Journal: Biomedical Reports

    Article Title: Evaluation of interferon-induced transmembrane protein-3 (IFITM3) rs7478728 and rs3888188 polymorphisms and the risk of pulmonary tuberculosis

    doi: 10.3892/br.2016.763

    Figure Lengend Snippet: IFITM3 rs3888188 T > G polymorphism using polymerase chain reaction-restriction fragment length polymorphism methods. The T allele was undigested (586-bp), while the G allele was digested by the Hpy 188I restriction enzyme and produces 340- and 246-bp

    Article Snippet: For the rs3888188 variant, the PCR products were digested with Hpy 188I restriction enzyme (Fermentas).

    Techniques: Polymerase Chain Reaction