Study On The Anther Culture And Microspore Culture In Wheat

In vitro anther culture and microspore culture were main ways to obtain haploids.Compared with the traditional breeding method, homozygosity is achieved in one generation,breeders can eliminate the numerous cycles of inbreeding required by conventional systems aswell as substantially reduce population sizes required for effective selection of superior traitcombinations. Assuming an efficient plant regeneration system is available, gametic cells arealso the preferred targets for transformation and transgenic studies.,The study of of antherculture for the production of wheat haploid plants is carried out earlier. Because anther cultureis technically simple, it is the main way to obtain wheat haploids at present.Only in recentyears has substantial progress been made in wheat doubled haploid production through theculturing of isolated microspores. Isolated microspore culture offers several advantages overanther culture. First, this method opens avenues for analyzing the basic mechanisms of pollenembryogenesis at the molecular and biochemical levels. Second, it also provides opportunitiesto increase the frequency of induction Thirdly, once freed from the barrier of anther walls,microspores become a favorable target for gene delivery, so it has been a hot area of researchfor the production of wheat haploid plants in recent years. Major factors affecting theefficiency of anther culture and microspore culture include genotype, donor plant physiology,microspore developmental stage, pretreatment conditions, physical and chemical conditionsfor induction culture, and plant regeneration. So It was of great significance that the researchwas to advance culture the efficiency of wheat anther and set up a high efficient culturetechnique system of the microspore culture of wheat.The research includes two aspects：on the one hand, it studies efficiency of anther culturefor different wheat genotype, sowing date and time of induction culture; on the other hand, itpreliminary studies wheat microspore culture. The main results are as follows:1. According to the experiment of Comparison of the anther culture response of16wheatgenotypes. The results indicate that the anther culture response was significantly differentamong genotypes, callus induction rate from0%to93.77%, green plant production rate from0%to7.92%when in the same culture conditions. The callus induction rates of Shi4185, Zhoumai11aiyouxi and Zhengyumai9987were higher respectively than other genotypes’,which were93.77%,47.33%and40.56%respectively. Meanwhile, Frequencies of greenseeding for all three genotypes were0.43%,0.67%and7.92%respectively. However, callusinduction rates of some genotypes were the lowest, which were all0, such as Zhengmai9962,Liangxing66, etc. According to the results, Zhengyumai9987was selected with high-cultureability.2. According to the experiment of comparison of efficiency of anther culture for differentsowing dates. The results indicate that sowing date is one of the significant factors of affectinganther culture. Although, on the second sowing date, which is15days later than the normalsowing date in Guanzhong, two materials anther culture, which were Fanmai8and Xinchun9,showed slightly lower at callus induction rate, showed the highest at frequency ofdifferentiation and differentiated plantlets. On the contrary, at the third sowing date, which is30days later than the normal sowing date in Guanzhong, two materials anther culture showedthat green plant production rates fell sharply from23.91%and19.57%which were themaximum value to0and2.25%respectively.3.According to the experiment of transfer of anthers (with and without embryo-likestructures) to regeneration conditions after a period of one to four weeks on induction medium(=anther-transfer treatment), The results indicate that callus induction rate of two materialsincreased with increased time, this value of induction culture after60days was11~12timesof induction culture after7days. Secondly, the rate of differentiated plantlets fell withincreased time. Finally, The rate of green seeding were the highest after7days on inductionmedium. So, transferring the anthers after7~14days caused a great improvement in theability of green plants to regenerate, which was conducive to differentiation of green plantlets.4. According to light microscopic observation of microspore development stages in theprocess of anther culture, The results indicate that the late-uninucleate microspores occurredplasmolysis by cold pretreatment (4℃,4d, for spike in vitro). Then, the size of embryonicmicrospore increased sharply by heat shock (32℃,3d, for excised anthers from cold-treatedspikes), and the star structures were clearly formed. Two cellular structures were clearlyformed after9days in culture. Multi-cellular structures were clearly formed after12days inculture with all cells still enclosed within the exine. Callus and embryoids were clearlyformed after28days in culture outside the exine.5. The research of microspore culture indicated that microspores after isolation andpurification didn’t bring about globular embryoids when they were cultured18days.