Development of four multiplex PCRs in the Zhikong scallop (Chlamys farreri) and their validation in parentage assignment
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In this study, three molecular assays (real-time multiplex polymerase chain reaction [PCR], merozoite surface antigen gene [ MSP ]-multiplex PCR, and the PlasmoNex Multiplex PCR Kit) have been developed for diagnosis of Plasmodium species. In total, 52 microscopy-positive and 20 malaria-negative samples were used in this study. We found that real-time multiplex PCR was the most sensitive for detecting P. falciparum and P. knowlesi . The MSP -multiplex PCR assay and the PlasmoNex Multiplex PCR Kit were equally sensitive for diagnosing P. knowlesi infection, whereas the PlasmoNex Multiplex PCR Kit and real-time multiplex PCR showed similar sensitivity for detecting P. vivax . The three molecular assays displayed 100% specificity for detecting malaria samples. We observed no significant differences between MSP -multiplex PCR and the PlasmoNex multiplex PCR kit (McNemar's test: P = 0.1489). However, significant differences were observed comparing real-time multiplex PCR with the PlasmoNex Multiplex PCR Kit (McNemar's test: P = 0.0044) or real-time multiplex PCR with MSP -multiplex PCR (McNemar's test: P = 0.0012).
Genetic algorithm
Plasmodium (life cycle)
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Objective To evaluate the application of fluorescent multiplex amplification of short tandem repeat(STR)for monitoring survival of engraftment after bone marrow transplantation(BMT).Methods Three STR loci named D12S391,D18S865 and D20S161 in 56 cases were detected by fluorescent multiplex amplification.PCR products were separated and typed by DNA Sequencer.Results The genotypes of STR in 52 recipient after bone marrow transplantation were completely identical with those of the donors.In another 4 cases the evidences of mixed chimerism were observed.Conclusion The system of fluorescent multiplex amplification of STRs exhibited high capacity of discrimination and low cost.Its application in the detection of STR after BMT is reliable,sensitive and simple.Combined with the clinical manifestation it can be used to evaluate the effect of BMT.
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Mariculture
Patinopecten yessoensis
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Patinopecten yessoensis
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Objective To establish a microsatellite-labeling multiplex PCR system of DNA for genetic quality monitoring of laboratory miniature pigs.Methods Three different fluorescence-modified microsatellite primers and ABI3700 genetic analyzer sequencing method were employed.After screening and optimizing the reaction conditions,a microsatellite-labeling multiplex PCR system of DNA for genetic quality monitoring of laboratory miniature pigs was established.Results Two multiplex PCR sets were optimized,with three microsatellite loci in each set.Genetic variation of three minipig populations were detected by the optimized primer sets.Conclusion The results indicate that the multiplex PCR system can provide a technical basis for studies on the genetic diversity and monitoring the genetic quality of inbred minipigs.
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Genetic monitoring
Primer (cosmetics)
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By simultaneously amplifying several loci in the same reaction, multiplex PCR has been used in gene mapping and DNA typing with polymorphic short tandem repeat loci. Previous studies have discussed in detail the various parameters and conditions that influence the quantity of individual products generated by multiplex PCR. In practice, when a primer pair fails to amplify in a multiplex PCR for some individuals, singleplex PCR is often employed as a supplement to amplify the primer pair. However, the reliability of this procedure is unknown. In this study, we used six primer pairs from ABI PRISMTM Linkage Mapping Set version 2 to perform multiplex and singleplex reactions. The fluorescence-labeled amplification products were separated and detected on ABI PRISM 310 Genetic Analyzer. We found that for the marker D1S468, multiplex and singleplex reactions for the majority of individuals yielded reactions of different sizes. Therefore, the potential size difference between multiplex and singleplex reactions needs to be investigated. This investigation is essential to employ multiplex PCR supplemented with singleplex PCR in gene mapping and DNA typing.
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Primer (cosmetics)
STR multiplex system
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We describe a sensitive, reliable and reproducible method, based on three multiplex PCR assays, for the rapid detection of seven common α‐thalassaemia deletions and one α‐globin gene triplication. The new assay detects the α 0 deletions – – SEA , – (α) 20.5 , – – MED , – – FIL and – – THAI in the first multiplex PCR, the second multiplex detects the –α 3.7 deletion and ααα anti3.7 variant, the third multiplex detects the –α 4.2 deletion. This simple multiplex method should greatly facilitate the genetic screening and molecular diagnosis of these determinants in populations where α‐thalassaemias are prevalent.
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