Abstract: 【Objective】 The objective of this study is to establish an efficient method of qualitative screening a few of transgenic-positive individuals from a large number of samples based on PCR technique and DNA pooling strategy.【Method】 In a case study, 480 samples were randomly numbered and then a virtual 3 dimensional pooling bulks of template DNA were constructed. Each of 480 samples was put into 5×96-well plates well by well. The row pools, X dimension pools, were the samples from same row number of all plates and therefore 8 (A, B, C, …… G, H) pools were made. The line pools, Y dimension pools, were the samples from same line number of all plates and therefore 12 (1, 2, 3, ……11, 12) pools were made. The plate pools, Z dimension pools, were the samples from the same plate and therefore 5 (I, II, III, IV, V) pools were made. The rationals of the design was used in order to archive high efficiency because the cross among 3 dimensions is unique along with high sensitivity and specificity be provided with PCR technique. The 500 ng•µL-1 positive samples were 96× diluted by using ddH2O or negative DNA, respectively. At the same time, 13 double blind samples of Bt176 events, Bt176 and Cry1Ab positive samples, were mixed in 480 samples. By screening the 25 pools, the candidate samples were detected and then screened them again. 【Result】 The results showed that the sensitivity and specificity of the detection were not affected. What’s more, this way exactly screened out 13 positive samples after screening of 25 pools and they were the positive 13 samples. 【Conclusion】 This method can be used for screening a few numbers of positive individuals from a large number of accessions. The work load will be reduced around 80% compared with conventional sample by sample method.

Key words: transgenic-positive individuals, screening, pooling, PCR, high efficiency