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18 results on '"Lapa, Sergey"'

13. Spoligotyping of the Mycobacterium tuberculosis complex using on-Chip PCR.

Author

Lapa, Sergey, Kuzmin, Alexey, Chernousova, Larisa, and Mikhailovich, Vladimir

Subjects
*POLYMERASE chain reaction, *MYCOBACTERIUM tuberculosis, *MYCOBACTERIA, *MYCOBACTERIUM bovis, *FLUORESCENCE, *DATA analysis
Abstract

Aims The aim of this study was to develop a rapid PCR-based method for spoligotyping of Mycobacteria in the microarray format and to compare it to conventional spoligotyping by hybridization. Methods and results The method employs the on-Chip PCR technique with primers specific for 43 spacers that separate direct repeats (DRs) in the DR region of mycobacterial DNA. The primers were immobilized on gel-based microarrays, and PCR was performed directly on the chips. The PCR fluorescence images were acquired and processed using a portable fluorescence analyzer equipped with dedicated software. Analysis takes 1.5–2 hours and can be carried out on clinical samples without additional handling. The analytical sensitivity of the method was 103 copies of target DNA. The spoligotyping results of 51 samples produced by the proposed method and by conventional reverse hybridization approach were in full concordance. Conclusions High throughput capacity, computerized data analysis, compact equipment, and reliable results make the on-Chip PCR an attractive alternative to intra- and interspecific spoligotyping of Mycobacterium tuberculosis complex bacteria. Significance and Impact of Study Fast microarray-based spoligotyping technique using on-Chip PCR was developed. [ABSTRACT FROM AUTHOR]

Published
2023
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16. Species-Level Identification of Orthopoxviruses with an Oligonucleotide Microchip

Author

Lapa, Sergey, primary, Mikheev, Maxim, additional, Shchelkunov, Sergei, additional, Mikhailovich, Vladimir, additional, Sobolev, Alexander, additional, Blinov, Vladimir, additional, Babkin, Igor, additional, Guskov, Alexander, additional, Sokunova, Elena, additional, Zasedatelev, Alexander, additional, Sandakhchiev, Lev, additional, and Mirzabekov, Andrei, additional

Published
2002
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17. Identification of Rifampin-Resistant Mycobacterium tuberculosis Strains by Hybridization, PCR, and Ligase Detection Reaction on Oligonucleotide Microchips

Author

Mikhailovich, Vladimir, primary, Lapa, Sergey, additional, Gryadunov, Dimitry, additional, Sobolev, Alexander, additional, Strizhkov, Boris, additional, Chernyh, Nikolai, additional, Skotnikova, Olga, additional, Irtuganova, Olga, additional, Moroz, Arkadii, additional, Litvinov, Vitalii, additional, Vladimirskii, Mikhail, additional, Perelman, Mikhail, additional, Chernousova, Larisa, additional, Erokhin, Vladislav, additional, Zasedatelev, Alexander, additional, and Mirzabekov, Andrei, additional

Published
2001
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18. Identification of Rifampin-Resistant Mycobacterium tuberculosisStrains by Hybridization, PCR, and Ligase Detection Reaction on Oligonucleotide Microchips

Author

Mikhailovich, Vladimir, Lapa, Sergey, Gryadunov, Dimitry, Sobolev, Alexander, Strizhkov, Boris, Chernyh, Nikolai, Skotnikova, Olga, Irtuganova, Olga, Moroz, Arkadii, Litvinov, Vitalii, Vladimirskii, Mikhail, Perelman, Mikhail, Chernousova, Larisa, Erokhin, Vladislav, Zasedatelev, Alexander, and Mirzabekov, Andrei

Abstract

ABSTRACTThree new molecular approaches were developed to identify drug-resistant strains of Mycobacterium tuberculosisusing biochips with oligonucleotides immobilized in polyacrylamide gel pads. These approaches are significantly faster than traditional bacteriological methods. All three approaches—hybridization, PCR, and ligase detection reaction—were designed to analyze an 81-bp fragment of the gene rpoBencoding the ß-subunit of RNA polymerase, where most known mutations of rifampin resistance are located. The call set for hybridization analysis consisted of 42 immobilized oligonucleotides and enabled us to identify 30 mutant variants of the rpoBgene within 24 h. These variants are found in 95% of all mutants whose rifampin resistance is caused by mutations in the 81-bp fragment. Using the second approach, allele-specific on-chip PCR, it was possible to directly identify mutations in clinical samples within 1.5 h. The third approach, on-chip ligase detection reaction, was sensitive enough to reveal rifampin-resistant strains in a model mixture containing 1% of resistant and 99% of susceptible bacteria. This level of sensitivity is comparable to that from the determination of M. tuberculosisdrug resistance by using standard bacteriological tests.

Published
2001
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