| The Pre-harvest Sprouting of wheat will reduces the yield and quality and leds to loss on quiet economy in wheat production. Basing on present genetic engineering, this objective for this study was to transfer Anti-TrxS from phalaris coerulescens into wheat cultivars to gain a new material with a characteristic of Pre-harvest Sprouting resistance. Four aspects were studied in this paper. Firstly, selecting the optimal wheat genotypes as receptors for gene transformation and optimize the culture system of immature embryo from wheat; Secondly, co-transferring Anti-TrxS and Bar into optimal receptors,immature embryos(calli) of Yumai18-64, by microprojectile bombardment; Thirdly, detecting and analyzing the Anti-TrxS gene in the T0 regenerated plants and T1～T4 progenies genetically;Fourthly, α-amylase activity and seed germination of the transgenic wheat and CK,related to the Anti-TrxS, were analysed。 The main results are as following:1. In order to select the optimal wheat genotype for gene transformation, 14 commercial wheat varieties from huanghuai wheat region were studied. The result showed that the difference among the genotypes was significant and Yumai18-64, 00zhong13, zhengxin991,and zhengxin992 had a higher tissue culture ability and were ideal receptors.2. In order to optimize the culture system of wheat immature embryos regeneration, immature embryos of Yumai18-64 and zheng9023 were taken as materials. The different concentration VB1 were supplemented into MS. Two weeks later,the frequency of callus formation and the weight of fresh calli in inducing media and the frequency of green primordial differentiation in differentiating media were investigated. Four weeks later the percentage of differentiation were investigated in the differentiating media. The results indicated 10mg/L VB1 supplemented into MS media can improve the embryogenic callus development and increase the bud differention frequency.3. The immature embryos from Yumai47,Yumai34,Zheng9023,Jimai2, Zhengxin991 and Yangao1 were desiccated for 0h,6h and 12h respectively before differentiation culture. The resulte showed desiccation can improved the bud differentiation frequency of calli and especially 12h desiccation. The differentiation frequency of calli was increased by 25% for Zheng9023 and 26.8% for Jimai2.This indicated the desiccation was a good way to improve bud differentiation of calli.4. Based on the results of optimization, the Anti-Trxs gene and Bar gene were co-transferred into the immature embryos(cali) from Yumai18-64 as receptors and 42 resistant plantlets of T0 generation were obtained from 2110 transferred embryos. Furthermore, 10d culture before bombardation and two shoot times can increased the regeneration frequency.5. 6 transgenic plants were gained from 42 vigorous regenerated plantlets and Anti-TrxS was integrated into the wheat genome by PCR and PCR-Southern analysis. There were 27 resistant plants in the 42 regenerated plantlets By PPT (150mg/L )screening. This result indicated bar gene was integrated and expressed in the wheat genome.6. PCR, nested-PCR and digestion analysis of the Anti-TrxS gene from T1,T2 and T3 plants showed the foreign Anti-TrxS gene was inherited to the progenies. Based on molecular assay and genetic analysis, in six tested 02TY18 transgenic lines, the ratios of PCR+/PCR- were 1:0 in 3 lines and 3:1,5:1, 1:1 in other 3 lines respectively and. in T1,T2andT3 transgenic lines of 01TY18,01TY70,00TY5 and 00T89,the segregation ratios of Anti-TrxS gene were 3:1 and less than 3:1. In 8 transgenic T4 lines of 00TY5 and 00T89 tested by PCR and PCR-Southern and analyzed with segregation ratios (X2), the ratios were 1:0 in 4 lines, 3:1 in 2 lines and more than 3:1 in other 2 lines.7. α-amylase activity and seed germination were investigated in the transgenic T1 seeds. The results showed the α-amylase activity and germination percentage were decreased in transgenic seeds comparing with CK.
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