Study On Inheritance And Molecular Biology Detection Of Interleukin-4 Gene In Progenies And Hybrid Generation Of Chinese Cabbage

In this research, we molecularly detected and genetically analyzed the three candidate transgenic Chinese cabbage varieties (named Habai2, Weibai6 and Xinshiji) through PPT, Kna resistance screening, PCR, PCR-Southern hybridization and ELISA and Western hybrid technology etc. which was harbored with interleukin-4 (il-4) gene transferred by pollen-tube pathway. Allowing il-4 gene cabbage offspring for molecular biological detection.meanwhile, il-4 gene in T₅, T₆, T₇ on behalf of the Chinese cabbage in the self expression of the transfer rule and expression were discussed.And progeny have been tested positive for hybridization between strains of different lines in order to improve positive rate and protein expression quantity. In addition, Positive phenotype of progeny plants, disease resistance was investigated.In this study, a total of sowing seeds of T5 generation of 608, the actual emergence of 421, we get 280 PPT resistant strains, in addition to the separation of PPT resistance are not found: non-resistant plants = 3:1 Mendelian segregation law. PCR detection of il-4 positive strains of 26, the positive rate was 6.18%.T6 generation of planting 1536, the actual emergence of 1108,which screened 1007 PPT resistance 356, the other 101 were analysed directly by PCR, In addition to the separation of PPT resistance except WeiBaiAH-3-1-5-23-10,WeiBaiAH-2-10-46-92-13,HaiBaiBH-1-5-4-12-21 ,XingShijiBL1-37-72-4-5, WeiBai AH1-447-49-1-33, the other plants do not meet resistance: non-resistant plants = 3:1 Mendelian segregation law.detected 44 positive strains, the positive rate of 3.97%.T₇ generation behalf of planting 918, the actual emergence of 580, of which 339 were screened by PPT resistance 135, the other 241 were analysed directly by PCR, PPT resistance to segregation laws of Mendelian segregation except HaiBaiAH3-1-5-23-2-2 is not have the plants. PCR detected 13 positive strains were obtained, the positive rate was only 2.24%.In Kna resistence screening process, T5 were planted on behalf of 772, the actual emergence of 378, positive strains screened Kna 226, Kna resistance to separation of the resistant plants were found: non-resistant plants = 3:1 Mendelian segregation law of the plant are: WeiBaiEL-1-3-121-11, WeiBai EL-1-3-47-26,WeiBai EL-1-3-121-12 WeiBai EL-1-3-121-1, WeiBai EL-1-3-47-44, WeiBai EL-1-3-47-24, WeiBai EL-1-3-3-4.PCR positive strains detected by 18 strains, and the positive rate was 4.76%. the separation of il-4, in addition to WeiBai EL-1-3-121-11, WeiBai EL-1-3-47-24 plants, the plants do not meet the number of positive: negative = 3:1 Mendelian plant number separation law.T₆ were planted on behalf of 787, the actual emergence of 673, of which 660 screened positive by Kna 192 strains, 13 strains of other direct PCR. In addition to resistance to separation of resistant plants all do not meet outside: non-resistant plants = 3:1 Mendelian segregation law.Positive strains were 11 strains detected by PCR; the positive rate was only 1.63%. F1 generation of hybrid seedlings under test 225, detected by direct PCR-positive strains of 5, the positive rate was 2.22%. This is far below the theoretical value.PCR detected as positive from a random sample of seven plants samples were PCR-Southern hybridization showed that 6 strains were seized samples were positive, indicating that il-4 still exist in the transformation generation. Five of them were Western blot, in which three showed hybridization bands; further proof of IL-4 has been stably inherited and expressed in generations. On including hybrids, including 11 samples were ELISA positive test results, WeiBaiAH1-447-4-2-10 against the total soluble protein content was 0.4% higher, hybrids of the TSP is 0.03%. Some samples'expression is lower.