Affinity capillary electrophoresis of DNA for detection of single-nucleotide polymorphisms and point mutations

2006 ◽  
Vol 1111 (2) ◽  
pp. 120-126 ◽  
Author(s):  
Kae Sato ◽  
Miwa Onoguchi ◽  
Kazuo Hosokawa ◽  
Mizuo Maeda
Keyword(s):  
Capillary Electrophoresis ◽  
Single Nucleotide Polymorphisms ◽  
Point Mutations ◽  
Nucleotide Polymorphisms ◽  
Affinity Capillary Electrophoresis ◽  
Single Nucleotide

scholarly journals Causal Role of Single Nucleotide Polymorphisms within themprFGene of Staphylococcus aureus in Daptomycin Resistance

2013 ◽  
Vol 57 (11) ◽  
pp. 5658-5664 ◽  
Author(s):  
Soo-Jin Yang ◽  
Nagendra N. Mishra ◽  
Aileen Rubio ◽  
Arnold S. Bayer
Keyword(s):  
Staphylococcus Aureus ◽  
Single Nucleotide Polymorphisms ◽  
Point Mutations ◽  
Nucleotide Polymorphisms ◽  
Wild Type ◽  
Single Nucleotide ◽  
Content Type ◽  
Reading Frame ◽  
A Charge

ABSTRACTSingle nucleotide polymorphisms (SNPs) within themprFopen reading frame (ORF) have been commonly observed in daptomycin-resistant (DAPr)Staphylococcus aureusstrains. Such SNPs are usually associated with a gain-in-function phenotype, in terms of either increased synthesis or enhanced translocation (flipping) of lysyl-phosphatidylglycerol (L-PG). However, it is unclear if suchmprFSNPs are causal in DAPrstrains or are merely a biomarker for this phenotype. In this study, we used an isogenic set ofS. aureusstrains: (i) Newman, (ii) its isogenic ΔmprFmutant, and (iii) several intransplasmid complementation constructs, expressing either a wild-type or point-mutated form of themprFORF cloned from two isogenic DAP-susceptible (DAPs)-DAPrstrain pairs (616-701 and MRSA11/11-REF2145). Complementation of the ΔmprFstrain with singly point-mutatedmprFgenes (mprFS295LormprFT345A) revealed that (i) individual and distinct point mutations within themprFORF can recapitulate phenotypes observed in donor strains (i.e., changes in DAP MICs, positive surface charge, and cell membrane phospholipid profiles) and (ii) these gain-in-function SNPs (i.e., enhanced L-PG synthesis) likely promote reduced DAP binding toS. aureusby a charge repulsion mechanism. Thus, for these two DAPrstrains, the definedmprFSNPs appear to be causally related to this phenotype.

scholarly journals Multiplex, fluorescent, solid-phase minisequencing for efficient screening of DNA sequence variation

1996 ◽  
Vol 42 (9) ◽  
pp. 1391-1397 ◽  
Author(s):  
T Pastinen ◽  
J Partanen ◽  
A C Syvänen
Keyword(s):  
Single Nucleotide Polymorphisms ◽  
Solid Phase ◽  
Point Mutations ◽  
Nucleotide Polymorphisms ◽  
Dna Templates ◽  
Single Nucleotide ◽  
Large Numbers ◽  
Dna Sequence Variation ◽  
Hla Dqa1 ◽  
Efficient Screening

Abstract We developed a multiplex, solid-phase minisequencing method to detect multiple single-nucleotide polymorphisms in an undivided sample. The amplified DNA templates are first captured on a manifold. Then, with multiple minisequencing primers of various sizes, single-nucleotide extension reactions are carried out simultaneously with fluorescently labeled dideoxynucleotides. The size of the extended product, determined by using a DNA sequencing instrument, defines the site of the polymorphisms, and the incorporated nucleotide gives the identity of the nucleotide at each site. HLA-DQA1 typing was used as a model system to evaluate the method. The DR2 subgroup of the HLA-DRB1 gene was typed along with the DQA1 gene to demonstrate the feasibility of the method in analyzing multiple genes at multiple sites simultaneously. The method is generally applicable for screening any single-nucleotide polymorphisms or point mutations, and its manifold format allows practical handling of large numbers of samples.

Single nucleotide polymorphism (SNP) markers for benzimidazole resistance in veterinary nematodes

Parasitology ◽  
2007 ◽  
Vol 134 (8) ◽  
pp. 1077-1086 ◽  
Author(s):  
G. VON SAMSON-HIMMELSTJERNA ◽  
W. J. BLACKHALL ◽  
J. S. McCARTHY ◽  
P. J. SKUCE
Keyword(s):  
Single Nucleotide Polymorphisms ◽  
Point Mutations ◽  
Building Blocks ◽  
Snp Markers ◽  
Nucleotide Polymorphisms ◽  
Nucleotide Polymorphism ◽  
Single Nucleotide ◽  
Principal Mechanism ◽  
Tubulin Isotype ◽  
Veterinary Importance

SUMMARYResistance to the benzimidazole class of anthelmintics in nematodes of veterinary importance has a long history. Research into the mechanisms responsible for this resistance is subsequently at a more advanced stage than for other classes of anthelmintics. The principal mechanism of resistance to benzimidazoles is likely to involve changes in the primary structure of β-tubulins, the building blocks of microtubules. Specifically, point mutations in the β-tubulin isotype 1 gene leading to amino acid substitutions in codons 167, 198, and 200 of the protein have been associated with resistance in nematodes. These single nucleotide polymorphisms offer a means of detecting the presence of resistance within populations. In this mini-review, we focus on the prevalence and importance of these polymorphisms in three groups of nematodes: trichostrongylids, cyathostomins, and hookworms. A brief overview of existing strategies for genotyping single nucleotide polymorphisms is also presented. The CARS initiative hopes to exploit these known polymorphisms to further our understanding of the phenomenon of BZ resistance.

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