MA Wenhua, MO Xiaoting, ZHANG Ying, BAI Xue, YANG Fan, ZHANG Jian, LI Wanshui, ZHAO Xingchun
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Objective To establish a porcine multiplex amplification system that was to be validated. Methods 12 porcine STRs (SW240, S0386, S0655, PigSTR4C, PigSTR5C, PigSTR7B, PigSTR11B, 387A12F, PigSTR13E, PigSTR14A, PigSTR15A, PigSTR17A) were selected to construct a fluorescent multiplex system plus the sexual marker Amelogenin. Through the serial completion of designing primers, labeling fluorescence dye and optimizing experiment conditions, a porcine multiplex amplification system was built up and to be validated. Results A multiplex PCR system, containing porcine 12 STR markers and an Amelogenin gender marker, has been successfully established. The most appropriate amplification conditions of the system are 10μL of amplification volume, 60℃ annealing temperature and 28 cycles. With the requirements met on precision, accuracy, peak height balance, stability, species specificity and polymorphism, the system is at lowest quantity of 0.125ng DNA to obtain complete STR genotyping, showing its cumulative discrimination power (CDP) of 0.999999999999961, the cumulative matching probability (CMP) of 3.9208×10-14 and the cumulative excluding probability of paternity (CEP) of 0.9952, respectively. Conclusions The established fluorescent multiplex system is capable of being used for porcine individual identification and paternity test in real cases.