Study On Abnormal Segregation Of Foreign Gene Bar In Herbicide-resistant Rice 9311HR

Herbicide-resistant transgenic rice has obvious superiority on improving and fast checkup seed purity in hybrid rice. Some herbicide-resistant transgenic rice varieties were obtained by particle bombardment, Agrobacterium tumefaciens and pollen-tube pathway method. However, transfer herbicide-resistant gene bar into other variety by cross and backcross is a simple and effective approach to getting new transgenic rice material resistant to herbicide"Basta". So, it is necessary to study the inheritance of foreign gene bar in the backcrossing progeny. In present experiment, the inheritance of foreign gene bar was studied by the tracking investigation of the herbicide resistance of generations crossed or backcrossed with bar transgenic rice, and more analysis have been made for the abnormal segregation of foreign gene bar in herbicide-resistant rice 9311HR. The main results obtained were as follows.1. By cross breeding, the herbicide-resistant gene bar from the donor, a genetically modified organism (GMO) HR obtained from introduction of bar into japonica rice Jingyin119 by biolistic method, was transferred into the breeding lines Yangdao 6(9311), E32, Yangdao 4, 2403, Suyunuo, Ninghui 1 and Zhenxiangjing. Some lines of bar transgenic rice F1 , F2, F3, BC1F1, BC1F2, BC1F3, BC2F1, BC2F2 and BC2F3 have been developed by continuous backcrossing and self-crossing. The inheritance of bar gene in different separate population was studied. The results showed that the segregation of herbicide resistance was accorded with the law of Mendelian heredity of a single dominant gene in japonica varieties such as 2403, Suyunuo, Ninghui 1 and Zhenxiangjing. However, abnormal segregation was observed in indica variety 9311. The herbicide resistance would segregate again with a ratio of 1:1 in selfcrossing progenies from herbicide-resistant plants.2. The effects of foreign gene bar on main agronomic traits, pollen fertility and pollen germination were studied in 9311HR which developed by backcrossing from 9311 and HR. Results indicated that the herbicide-resistant gene bar had no significant effect on the main agronomic traits of herbicide-resistant plants of 9311HR. The pollen fertility and in vitro pollen germination rate for herbicide-resistant plants of 9311HR were about 90%. Also, the herbicide-resistant plants of 9311HR showed normal seed fertility. There existed no siginificant difference in pollen fertility, in vitro pollen germination rate and seed fertility between 9311HR and 9311. Moreove, PCR analysis of pollen DNA indicated that mature pollen which carrying foreign gene bar were formed in herbicide-resistant plants of 9311HR. Above results sugested that there was no gametal sterility in herbicide resistant plants of 9311HR.3. In order to know the heredity transmission of bar gene in the progeny, the herbicide-resistant plant segregated from 9311HR was used as female or male to cross with conventional rice variety Zhenxian 96 or (HR-2×881)F9 or CMS line, and the segregation ratio of herbicide-resistant to sensitive plants was investigated in the test F1 poppulation. When the resistant plant was used as female parent, the segregation ratio of resistant to sensitive plants was 1:1. However, when it was used as male parent, all the progenies were sensitive to herbicide. Result of PCR analysis indicated that bar gene amplification of the herbicide resistant plants were positive and the herbicide sensitive plants were negative. The biological and genetic basis of distorted segregation of bar gene was dicussed and deduced on the basis of above results that foreign gene bar only could be transferred to progenies by pistils. The authors are inclined to ascribe these phenomena to the absent impregnation of japonica pollen which carried herbicide-resistant gene bar.4. To genetically map the foreign bar , 368 F2 plants developed by crossing 9311 with HR were used as mapping population. Genomic DNA from F2 plants of 9311×HR was extracted and analyzed for co-segregation, using available molecular markers. Mapping results showed that the foreign gene bar was located between SSR marker RM5551 and RM6080 at a distance of 4.4cM from RM5551 and 1.2cM from RM6080 on chromosome 3. We presumed that foreign gene bar was tightly linked with gametophyte gene of ga2 on chromosome 3. It conduced gametophytic sterility of japonica male gametes which carried foreign gene bar. Thereby, it caused abnormal segregation of herbicide resistance in 9311HR.