Selection Of The F₂ Recombinant Plants In Five Chromosomal Segment Substitution Lines And The Selection And Cloning Of The Herbicide HW-02 Resistant Gene

There are many factors impacting the plant tissue culture, including plant genotype. In general the monocotyledon is more difficult than dicotyledon in tissue culture. A high efficient tissue culture system has been established and applicated in japonica rice which is considered as the model variety in the field of plant tissue culture. However, the tissue culture in most of indica rice varieties is still difficult,and genotype is a major obstacle.Some quantitative trait loci(QTLs) related to frequency of callus induction,efficiency of callus subculture,frequency of plant regeneration were mapped by using a chromosomal segment substitution lines(CSSL) population derived from a cross between an indica rice variety "Zhenshan 97B" and japonica rice variety "Nipponbare".According to the results made by Zhao (Zhao et al, 2009),five CSSL lines HJ-36, HJ-37, HJ-47, HI-89 and HJ-119 were selected and backcrossed with Zhenshan 97B in Hainan province. Sixty eight alternative SSR(simple sequence repeat) markers were screened for the detection of the recombinant plants and eight SSR markers showed specificity between the parent lines. Thity one recombinant plants were obtained in HJ-37 F₂ population, twenty nine recombinant plants were obtained in HJ-47 F₂ population, twenty four recombinant plants were obtained in HJ-89 F₂ population and thirty two recombinant plants were obtained in HJ-119 F₂ population. Twenty out of twenty seven SSR markers were selected for the fine mapping of the four CSSLs, among which four SSR markers were used for the fine mapping of HJ-37 F₂ population, four for HJ-47, six for HJ-89, and six for HJ-119.HW-02 is a novel type ofα-(substitutted phenoxy acetoxy) alkyl phosphonates chlorophenoxy herbicide which targets pyruvate decarboxylase. We have confirmed that the DH10B strain is much more sensitive to the herbicide than DH5αstrain. In this study we obtain two herbicide-resistant DH5αmutants by EMS (Ethyl Methane Sulfonate) mutagensis. The pyruvate decarboxylase gene from the mutants was cloned by PCR and constructed on the expression vector. Then we introduce the expressiom vector into DH10B to express the protein.