bsmai  (New England Biolabs)

Journal: American journal of medical genetics. Part A

Article Title: Somatic Mosaicism in Menkes Disease Suggests Choroid Plexus-mediated Copper Transport to the Developing Brain

doi: 10.1002/ajmg.a.33632

Figure Lengend Snippet: Analysis of mosaicism. A. Diagram of ATP7A exon 15 region that was amplified from genomic DNA from the mosaic Menkes disease patient and a normal control, showing position of a novel restriction site ( BsmAI ) created by the P1001L mutation. The 294 bp

Article Snippet: The 294 bp PCR products generated were purified using a DNA isolation kit (QIAGEN, Valencia, CA, USA), digested with BsmAI (NEB, Beverly, MA) and electrophoresed through 2% agarose.

Techniques: Amplification, Mutagenesis

Journal: Journal of Cellular and Molecular Medicine

Article Title: Pathogenicity of two COQ7 mutations and responses to 2,4‐dihydroxybenzoate bypass treatment

doi: 10.1111/jcmm.13154

Figure Lengend Snippet: COQ7 gene variants and a mitochondrial DNA mutation identified in the patient under study. ( A ) COQ7 gene variants. Sequencing chromatograms are shown for the c.308C > T and c.332T > C variants detected in COQ7 in the proband reported in this study, with the wild‐type sequence at the top. The chromatograms also show that the patient is homozygous for both the c.308C > T [p. Thr103Met] polymorphism and the c.332T > C [p.Leu111Pro] mutation. ( B ) Chromatograms showing a partial sequence of the mitochondria 12S rRNA gene in a normal control, the L111P patient reported in this study or the previously reported COQ7 patient carrying the V141E mutation. Both patients and the normal control used in this study carry the m.1438A > G polymorphism. The L111P patient, in addition, carries the m.1555A > G mutation. Black arrows denote altered nucleotides. ( C ) RFLP of mitochondrial DNA. Shown is an agarose gel electrophoresis of PCR products from the mitochondrial 12S rRNA gene digested by BsmAI. Wild‐type control: skin fibroblasts from a healthy unrelated individual; L111P: the proband reported in this study; V141E: COQ7 patient previously reported in 19 . The PCR product (643 bp) without the m.1555A > G mutation was cleaved in two fragments of 409 and 234 bp, which was the case for the wild‐type control and the V141E patient. The PCR product from the L111P patient's 12S rRNA gene was not cut by BsmAI, as the m.1555A > G mutation abolishes the BsmAI restriction site. Moreover, only a single band corresponding to the mutated fragment was seen in the sample from the L111P patient. This, together with the sequencing chromatogram, demonstrates homoplasmy for the m.1555A > G mutation.

Article Snippet: The 643‐bp PCR product was subjected to sequencing using the same primers as well as restriction fragment length polymorphism (RFLP) analysis using BsmAI (New England Biolabs).

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

Journal: The Journal of Neuroscience

Article Title: Slow-Channel Myasthenic Syndrome Caused By Enhanced Activation, Desensitization, and Agonist Binding Affinity Attributable to Mutation in the M2 Domain of the Acetylcholine Receptor α Subunit

doi: 10.1523/JNEUROSCI.17-15-05651.1997

Figure Lengend Snippet: Identification and analysis of mutation in the AChR α subunit. A , Automated sequencing of α exon 7 around codon 249 in a control and the propositus. In the propositus, both G and T nucleotides are present at position 745 ( arrow ), indicating a heterozygous G→T transversion. This mutation changes codon 249 from a GTC for valine to a TTC for phenylalanine. B , Allele-specific PCR and mutagenic PCR plus Bsm AI analysis of genomic DNA in the propositus’ family. Both wild-type and mutant-allele-specific primers amplify an expected 168 bp fragment in propositus and his father, but only the wild-type primer amplifies the expected fragment in the other family members. On Bsm AI analysis after mutagenic PCR, the wild-type allele gives rise to a 134 bp fragment ( open arrowhead ) and a 23 bp fragment (not shown); the mutant allele yields an undigested 157 bp fragment ( closed arrowhead ). Both wild-type and mutant fragments appear in the propositus, but only the wild-type fragment appears in other family members. The incongruity between allele-specific PCR and restriction analysis in the father is attributable to the father being a mosaic for αV249F (see text). The arrow indicates propositus. C , Multiple alignment of AChR M2 membrane-spanning domains. The boxes enclose the conserved valine residues in human AChR subunits and in AChR α subunits of other species. The mutant phenylalanine is shown at the bottom.

Article Snippet: The PCR product was digested with Bsm AI (New England Biolabs, Beverly, MA).

Techniques: Mutagenesis, Sequencing, Polymerase Chain Reaction

Journal: Plant Physiology

Article Title: The lss Supernodulation Mutant of Medicago truncatula Reduces Expression of the SUNN Gene 1 Gene 1 [OA]

doi: 10.1104/pp.110.164889

Figure Lengend Snippet: Origin of expressed SUNN in sunn LSS/ lss SUNN plants. A, Diagram of mutation and PCR product locations and restriction sites used in B. H indicates a Hae III restriction site, while B indicates a Bsm AI site; within the PCR product, differential digestion

Article Snippet: Following amplification, the PCR products were digested with Bsm AI (New England Biolabs) at 55°C for 4 h, which cuts only the sunn-2 allele, or Hae III (New England Biolabs) at 37°C for 4 h, which cuts only the sunn-1 allele.

Techniques: Mutagenesis, Polymerase Chain Reaction