Abstract: Bovine leukocyte adhesion deficiency (BLAD) was an autosomal recessive lethal genetic disease, the caused BLAD had a bad effect on the bovine production trait and dairy economic returns. In order to establish a rapid and reliable method for BLAD detection, in this experiment, according to BLAD CD18 gene sequences published in GenBank, PCR primers were designed. After PCR amplification and purification, three standard plasmids including genotypes A/A, A/G and G/G were constructed. Pyrosequencing method were established by using these constructed standard plasmids. Then, the bovine blood samples were detected by the method. The results were compared with Sanger sequencing results to analyze and verify the accuracy of the test results. 300 bovine blood samples of BLAD were detected by pyrosequencing method，the results showed A/A genotype were 294 cases, accounting for 98% of the overall proportion; A/G genotype were 6 cases, accounting for 2%; and the results of two detection methods were consistent. It showed that pyrosequencing method to detect BLAD were specific, sensitive and rapid, and the test results were accurate and reliable. It was suitable for kit development and applied in clinical diagnosis of BLAD.

Key words: font-size: 10.5pt; mso-bidi-font-size: 12.0pt; mso-font-kerning: 1.0pt; mso-bidi-font-family: 'Times New Roman'; mso-ansi-language: EN-US; mso-fareast-language: ZH-CN; mso-bidi-language: AR-SA" lang="EN-US">bovine leukocyte adhesion deficiency; CD18 gene; pyrosequencing method