© The Korean Society for Laboratory Medicine. Identification of raw and heat-processed meats from game bird species by polymerase chain reaction-restriction fragment length polymorphism of the mitochondrial D-loop region. baumannii gyrA and parC mutations with FQ resistance.Īcinetobacter baumannii Fluoroquinolone resistance PCR-restriction fragment length polymorphism Quinolone resistance-determining regions. Study with Quizlet and memorize flashcards containing terms like What are restriction endonucleases, What do restriction endonucleases do, Why will the length of cut DNA fragments vary between individuals and species and more. Journal of Agricultural and Food Chemistry 2007, 55 (24), 9913-9920. This unit describes Restriction Fragment Length Polymorphism (RFLP) analysis, which utilizes restriction endonuclease digestion to identify DNA sequence polymorphisms in genes or DNA regions of interest. This assay specifically amplified gyrA and parC from A. As for the non- baumannii Acinetobacter strains, although amplification products for gyrA were obtained for 28 strains, no parC amplification product was obtained for any strain. PCR products from 47 FQ-resistant strains containing mutations in gyrA and parC were not digested by the restriction enzymes at the site containing the mutation. In 11 FQ -susceptible strains, the gyrA and parC PCR products were digested by the selected restriction enzymes at the site containing gyrA (codons 83 and 87) and parC (codons 80 and 84). PCR amplification of gyrA and parC was successful for all A. Restriction endonuclease analysis based on sequence data of those fragments was used for diffferentiation among species. baumannii strains and 37 other Acinetobacter strains that have been identified by RNA polymerase β-subunit gene sequence analysis. baumannii gyrA and parC, two primer sets were designed for mismatched PCR-RFLP to detect mutations in gyrA (codons 83 and 87) and parC (codons 80 and 84) by introducing an artificial restriction enzyme cleavage site into the PCR products.
baumannii.īased on the conserved sequences of A. We developed a mismatched PCR-restriction fragment length polymorphism (RFLP) assay to detect mutations in the gyrA and parC QRDRs associated with FQ resistance in A. Thermo Fisher Scientific dyes are available in multiple chemical forms. The RFLP polymorphism is caused by a C-T transition at nucleotide 421 on the cDNA, resulting in a threonine-to-isoleucine conversion. Mutations in the quinolone resistance-determining regions (QRDRs) of Acinetobacter baumannii DNA gyrase ( gyrA) and topoisomerase IV ( parC) are linked to fluoroquinolone (FQ) resistance. What is a Restriction Fragment Length Polymorphism (RFLP) After DNA was confirmed as the hereditary genetic material, the burgeoning field of molecular biology exploded with a multitude of. Terminal restriction fragment length polymorphism (T-RFLP).
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