Abstract: It’s important for separation offspring in transgenic Bt gene broccoli to identify positive plant lines and resistant materials accurately by different methods. In the study，five backcross materials based on four to five generations，one homozygous material as positive control and five inbred lines as negative controls were used for analysis of the insect-resistant effectiveness of Cry1Ac in transgenic broccoli from the levels of gene，protein and appearance. The gene of Cry1Ac was carried out by PCR amplification，protein was carried out by determination of Bt-Cry1Ab/1Ac test strip，and appearance was carried out by feeding test of diamondback moth in vitro. At the same time，the advantages of three methods were evaluated and stated. The result showed that a total of 82，77 and 75 positive strains were divided from 159 strains of five backcross materials. According to chi-square test at the level of 5%，the positive and none positive strains of 1︰1 were calculated and in accordance with mendelian inheritance. By comparisons of three methods，we found that the feeding trial of diamondback moth in vitro was accurate and reliable，but the experimental conditions must to be well controlled. The Bt-Cry1Ab/1Ac test strip was sensitive，fast and can observe the actual Bt protein expression，but it must control human errors. The normal PCR amplification could be used to identify the positive strains，but it can not reflect the actual anti-insect effect，so it needs to be together with the test of feeding diamondback moth or the test of Bt protein，which could provide an accurate result.

Key words: broccoli, transgene, Cry1Ac, diamondback moth, heridity