| As a new discovery in wheat breeding in the 20 th century,the photo-thermo-sensitive male sterility in wheat provides a new breakthrough for the use of wheat heterosis and the improvement of global wheat production.The study on the regulation of the genes related to wheat photo-thermo-sensitive male sterility and the genetic mechanism of infertility has become a hot topic in the field of wheat male sterility.On the basis of previous research in our laboratory,this study obtained a number of genes related to reproductive development from the photo-thermo-sensitive male sterility line 337 S,screened out differentially expressed gene,and cloned it as candidate gene for this study.This study used Gateway technology to construct the overexpression vectors of candidate genes(TaMS337S-1,TaMS337S-3 and TaMS337S-4),and introduced it into different wheat cultivars by means of gene bombardment and Agrobacterium tumefaciens to verify gene function,aimed to explore the effects of the expression of three candidate genes on the fertility of wheat and its role in the process of infertility regulation.Also,it provide help for the resolution of sterility mechanism of the photo-thermo-sensitive male sterility line 337 S.The main results are as follows:1.Agrobacterium-mediated genetic transformation of immature embryos: There were 4 strains of resistant seedlings obtained from the TaMS337S-3 gene transformation experiment which uses BobWhite as receptor material,and they were all negative by PCR test.And we obtained 17 strains of resistant seedlings through the TaMS337S-1 gene transformation experiment which uses Huamai 2566,Huamai 2152,Huamai 1168,Huamai 1223 and Longchun 23 as receptor material.It can be preliminarily concluded that there were 6 positive plants including 2 plants of Huamai 2566,1 plant of Huamai 2152,1 plant of Huamai 1168 and 2 plants of Longchun 23,but Huamai 1223 did not get resistant seedlings.2.Transformation of immature embryos by gene gun: We got 54 strains of TaMS337S-1 transgenic resistant seedlings which include 7 plants of Longchun 23(3 positive seedlings),40 plants of Huamai 2566(12 positive seedlings),7 plants of BobWhite(2 positive seedlings).And there were 40 strains of TaMS337S-3 transgenic resistant seedlings including 5 plants of Huamai 2566(only one was positive),19 plants of BobWhite(5 positive seedlings),16 plants of Kenong 199(3 positive seedlings).The ratio of resistant seedlings and positive seedlings of TaMS337S-4 transgenic regrowth: Kenong 199 13: 4,BobWhite 7: 3.3.Transformation of mature embryos by gene gun: Huamai 1223 and Huamai 1168 were bombarded with the bombardment pressure of 900 psi and 650 psi and the bombardment distance of 9 cm.The comparison showed that the ability of differentiation and regeneration of the callus under the bombardment pressure of 900 psi was stronger than 650 psi.And we initially got 4 strains of resistant Huamai 1168 seedlings under the bombardment pressure of 900 psi in the transformation experiment of TaMS337S-4 gene,but no positive seedlings were obtained finally.4.The phenotypic characteristics of transgenic positive seedlings: Compared with the corresponding recipient parents,the positive transgenic plants transfected with TaMS337S-4 gene are generally short and weak.Most plants only have one ear,and the ear is thin,some ears even appears phenomenon of insufficient grouting.The positive seedlings transfected with TaMS337S-3 gene showed vigorous vegetative growth and delayed reproductive growth.The specific performance was as follows: transgenic plants continuously tilted,the heading time delayed about 40 days,and the number of headings raised with the increase of tillers number. |
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