scholarly journals Polymerase chain reaction primer sets for the detection of genetically diverse human sapoviruses

2020 ◽  
Vol 165 (10) ◽  
pp. 2335-2340
Author(s):  
Tomoichiro Oka ◽  
Seiji P. Yamamoto ◽  
Nobuhiro Iritani ◽  
Shigenori Sato ◽  
Chika Tatsumi ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Polymerase Chain Reaction Primer ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Primer Sets

Novel subgingival bacterial phylotypes detected using multiple universal polymerase chain reaction primer sets

2006 ◽  
Vol 21 (1) ◽  
pp. 61-68 ◽  
Author(s):  
A. Lillo ◽  
F. P. Ashley ◽  
R. M. Palmer ◽  
M. A. Munson ◽  
L. Kyriacou ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Polymerase Chain Reaction Primer ◽  
Chain Reaction ◽  
Bacterial Phylotypes ◽  
Polymerase Chain ◽  
Primer Sets

scholarly journals Novel multiplex polymerase chain reaction primer set for identification of Lactococcus species

2011 ◽  
Vol 52 (5) ◽  
pp. 491-496 ◽  
Author(s):  
T. Odamaki ◽  
S. Yonezawa ◽  
M. Kitahara ◽  
Y. Sugahara ◽  
J.-Z. Xiao ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Multiplex Polymerase Chain Reaction ◽  
Polymerase Chain Reaction Primer ◽  
Chain Reaction ◽  
Polymerase Chain

scholarly journals Genotyping of Listeria by Polymerase Chain Reaction (PCR) and its Epidemiological Significance

2021 ◽  
Vol 6 (2) ◽  
pp. 115-124
Author(s):  
V. Zyuzin ◽  
◽  
O. Tuzova ◽  
U. Frenkel ◽  
Muntian L. ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Internal Control ◽  
Pcr Analysis ◽  
Chain Reaction ◽  
Virulence Markers ◽  
Chip Technology ◽  
Polymerase Chain ◽  
Primer Sets ◽  
Epidemiological Significance

The purpose of the study. The article covers the issues of genotyping of listeria by polymerase chain reaction (PCR) and its epidemiological significance. It is known that molecular genetic methods allow to detect specific microbial pathogens, virulence markers, antimicrobial resistance genes faster and with greater sensitivity than traditional culture methods. Therefore, the development of detection methods and genotyping by polymerase chain reaction (PCR) is relevant. Materials and methods. For the detection and genotyping of Listeria, the technology of DNA chips is becoming increasingly important, which can significantly expand the possibilities of molecular detection. Chip technology can be used to simultaneously identify a whole range of pathogenic microorganisms, to determine genetic virulence markers, the relationship to antibiotics, subtyping, as well as to determine the quality of microorganisms in samples. A simplified version of DNA chip technology is multiplex (numerical) PCR, which is used to detect and genotype listeria. Studies have shown that to detect Listeria spp. using a polymerase chain reaction, it is advisable to use the gene iap (invasive associated protein), known for 6 species of listeria, which encodes a protein P 60 that is common to all species of listeria, including L.murrayi. Computer analysis revealed areas with 100% homology, from which primers were selected for PCR detection of all types of listeria. Areas of genomes characterized by 100% homology were selected for further analysis and labeling of primer sets. The sequences of the constructed primers List 1 and List 2 allowed to identify 6 species of Listeria (L. monocytogenes, L. innocua, L. ivanovii, L. grayi, L. seeligeri, L. welshimeri). Increasing the length of the primer leads to the increasing of specificity of PCR analysis. The greater the length of the primer, the smaller the specific gravity of one error of the unpaired nucleotide. The degree of primer homology is a key parameter that indicates the "quality" of a set of primers. Results and discussion. It is established that a significant disadvantage of the vast majority diagnosed using PCR test systems is the lack of internal control of amplification. The negative result of PCR analysis may be due to the absence in the clinical material of a fragment of the Listeria genome, and the fact that the PCR product was not synthesized for other reasons. They may be as the following ones: operator errors, erroneously determined reaction mixture concentrations and PCR temperature parameters. False-negative results can also be caused by factors that inhibit thermostable DNA polymerase. In its turn, such inhibition of the enzyme responsible for amplification is caused by a very large amount of DNA - template, pre-treatment of clinical samples. It has been shown that 80% of clinical specimens contain a substance that inhibits DNA polymerase. Therefore, it is necessary to use internal control, the positive result of the reaction of which indicates the successful amplification, that is the absence of false positive results. Conclusion. There are several reasons why the accuracy of PCR analysis does not reach 100%. Accuracy depends on the technology (variety) of PCR - the method used (ordinary or fluorescent), detection of amplicons, PCR homogeneous or nested, nested in one test tube or in two test tubes, as well as the level of quality of the survey (primarily on the technical parameters of the amplifier). The test systems used can be used for PCR detection and are recommended as standard primer sets for the detection and cross-species testing of listeria, which is important for the timely implementation of appropriate anti-epidemic measures in listeriosis

scholarly journals Prevalence of the Helicobacter pylori babA2 Gene in Children Mainly Depends on the PCR Primer Set Used

2020 ◽  
Vol 2020 ◽  
pp. 1-9
Author(s):  
Anja Šterbenc ◽  
Maja M. Lunar ◽  
Matjaž Homan ◽  
Boštjan Luzar ◽  
Nina Zidar ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Helicobacter Pylori ◽  
Diagnostic Tool ◽  
Clinical Relevance ◽  
Detection Methods ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Primer Sets ◽  
H Pylori ◽  
Pcr Primer

Various polymerase chain reaction- (PCR-) based methods with varying positivity rates were designed to detect the Helicobacter pylori babA2 gene. To compare different primer sets, babA2 prevalence was determined in 279 H. pylori-positive pediatric samples using the 832 bp, 139 bp, and 271 bp PCR primer sets, resulting in 34.0%, 51.3%, and 79.6% prevalence of the babA2 gene, respectively. The babA2 status determined using the 832 bp and 139 bp PCR primer sets significantly correlated with bacterial density and activity of inflammation, whereas no such correlations were found using the 271 bp PCR primer set. The 139 and 832 bp PCR primer sets concordantly detected the babA2 gene in 93 cases; however, in comparison to the 832 bp PCR primer set, the 139 bp PCR primer set detected additional 50 babA2 cases, whereas only two 832 bp positive cases were missed. The 271 bp PCR primer set missed 32 babA2 cases that were 832 bp and/or 139 bp PCR positive, but tested solely positive in 109 cases. Interestingly, cloning of a subset of 271 bp PCR positive samples revealed amplification of the babA/B gene chimera. Hence, in our opinion, the 271 bp PCR protocol is not a reliable diagnostic tool for detecting the babA2 gene in children. Our results reaffirm previous observations that the use of certain babA2 PCR primer sets can significantly impact estimation of the prevalence and clinical relevance of the H. pylori babA2 gene in children, suggesting babA2 detection methods should be carefully selected.

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