牛先天性下痢致死基因與單譜症基因同時檢測的方法

林德育 黃鈺嘉 陳若菁 張秀鑾

行政院農業委員會畜產試驗所

建立多種基因同時檢測的方法不僅可節省檢測時間與人力,亦可降低檢測成本。本試驗嘗試利用Mutiplex PCR針對牛淋巴球黏力缺失症(Bovine Leukocyte Adhesion Deficiency,曾譯為牛遺傳性白血症,BLAD)與牛單譜症(Deficiency of Uridine Monophosphate Synthase,直譯為尿核F單磷酸鹽合成@缺失症,國際上簡稱為 DUMPS)兩個重要的遺傳疾病開發同時檢測的方法。以已知檢測牛淋巴球黏力缺失症引子與牛單譜症的引子同時進行PCR增幅不同特定DNA片段,再同時以限制@TaqI與AvaI依序以37℃ 3小時與65℃ 4 小時進行分切,分切後產物以4% Agarose電泳分析。由於所得的DNA片段長度有明顯的差異,所以易於判讀不同的遺傳型,顯示本方法確實可同時檢測此兩種遺傳疾病。

關鍵語:多樣的聚合@連鎖反應、牛淋巴球黏力缺失症、單譜症。

 

SIMULTANEOUS ANALYSIS OF BOVINE BLAD AND DUMPS ALLELES BY
MULTIPLEX PCR FOLLOWED BY DIGESTION WITH TWO RESTRICTION ENZYMES

D. Y. Lin, Y. C. Huang, Z. C. Chen and H. L. Chang

Taiwan Livestock Research Institute, Council of Agriculture

An improved and simplified method allowing simultaneous genetic typing of Bovine Leukocyte Adhesion Deficiency(BLAD) and Deficiency of Uridine Monophosphate Synthase(DUMPS) loci has been developed. The method is based on simultaneously amplified the fragments of two genetic disease alleles by multiplex PCR, and concurrently digested the products by two restriction enzymes(TaqI and AvaI mixed together; 37℃ 3hr and then 65℃ 4hr) in the same buffer. All combinations of the known normal and mutant alleles could be detected by electrophoretic separtion performed on the same agarous gel owing to the obvious differences in the length of the restriction fragments. The expected benefits of increased speed and decreased labour and cost seem to be worthwhile to use this improved method.

Key Words: Multiplex PCR, BLAD, DUMPS.