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《Journal of Agricultural Biotechnology》 2003-03
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Establishment of the Method for Identifying the Extracellular Fatty Acid Binding Protein Genotypes with Allele-specific-PCR

Zhao Chunjiang Li Ning* Deng Xuemei (State Key Laboratory for Agrobiotechnology, China Agricultural University, Beijing 100094, China)  
The genotypes of chick (Gallus gallus ) extracellular fatty acid binding protein (EX-FABP) are closely related to chick abdominal fat percentage. In the study, the method for identifying the EX-FABP genotypes with allele-specific PCR (AS-PCR) technique was established and the strategy of identifying single nucleotide polymorphisms with AS-PCR was discussed. A typical AS-PCR system consists of two allele-specific primers and ane common primer. PCR products can be resulted when a sample's DNA is amplified by the common primer and allele-specific primer which matches with the sequence of sample's DNA. There are not PCR products if the allele-specific primer does not match the sequence of the samples' DNA. According to whether there is PCR products or not, the genotypes of samples can be identified. For establishing an AS-PCR, it is very important to design proper number and kinds of mismatched bases at the 3′ end of the allele-specific primers where the mutation occurs and different genotypes are generated. When one mismatched base in the primers can not lead to different PCR results which can identify genotypes, the second or even the third mismatched bases should be added to the allele-specific primers. The different kinds of mismatched bases have different effect on the results of AS-PCR. If the mismatched bases are A-C or T-G, they have little effect on the amplifying efficiency of AS-PCR. But when the mismatched bases are A-G or C-C or C-G, they have dramatically negative effect on it. A modified AS-PCR, allele-specific and length-deferent PCR (ASLD-PCR) were established and introduced. In an ASLD-PCR system, two pairs of primer sites, each of them consists of an allele specific primer and a common primer,were used to amplify and identify genotypes. The PCR products of the two pairs of primers were different in the length and the different length of PCR products standed for different kinds of genotypes. So ASLD-PCR was more convenient to identify genotypes than AS-PCR.
【CateGory Index】: S831
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