【作者】 唐宗祥；

【导师】 任正隆；

【作者基本信息】 四川农业大学 ， 生物化学与分子生物学， 2003， 硕士

【摘要】 本研究采用四川省重点推广的小麦新品种R57(川农17)和R59(川农12)的成熟胚及R59的去胚芽成熟胚为外植体，进行离体培养研究，以建立高效的小麦再生体系，从而解决小麦组织培养过程中外植体来源受季节限制的问题，并为进一步进行小麦转基因研究打下基础。同时也将两个品种的组织培养特性与Bobwhite的组织培养特性作比较。现将本研究结果概括如下： 1．本研究建立了以小麦成熟胚为外植体的高效适用的小麦植株再生体系，即以完整成熟胚为外植体，在诱导培养基中添加2mg/l 2,4-D，将经过14天诱导的愈伤组织，于附加5mg／lKT的分化培养基中进行分化培养，这样的组合能取得最好的效果。 2．R57、R59两个品种的成熟胚有相似的优良组织培养特性，其成愈率和愈伤组织分化率在同类研究中位居前列。在不同诱导培养基中，R.57成熟胚平均成愈率最低为77.28％，最高达97.58％。R59成熟胚最低平均成愈率为79.22％，最高为99.33％。在最适宜的培养条件下，R57愈伤组织平均分化率为73.63％。R59愈伤组织平均分化率可达89.23％。 3．种子的收获时期和培养前的低温处理对成熟胚愈伤组织的诱导效应有影响。2002年收获的且经低温处理的成熟胚比2001年收获的未经低温处理的成熟胚有更高的成愈率，且在大多数诱导培养基中差异显著。2001年的成熟胚在不同诱导培养基中的成愈率存在显著差异，而2002年的成熟胚在不同诱导培养基中的成愈率差异不显著。 4．本文首次报道了种子的收获时期和培养前的低温处理对愈伤组织诱导过程中成熟胚萌发成苗的影响。2002年的成熟胚比2001的成熟胚有更高的发芽率，且差异极显著。本研究还表明，高浓度2,4-D有抑制成熟胚萌发的作用，KT有促进成熟胚萌发的作用。 5．本文首次报道了胚芽对成熟胚愈伤组织的形成及愈伤组织的分化能力的影响。胚芽对愈伤组织的形成和愈伤组织的分化都有利。R59去胚芽成熟胚的成愈率低于完整成熟胚的成愈率，且在多数诱导培养基中存在显著或极显著差异。Bobwhite去胚芽成熟胚的成愈率和完整成熟胚的成愈率无差异，都达100％。然而，完整成熟胚来源的愈伤组织比去胚芽成熟胚来源的愈伤组织分化率更高，且两者存在极显著差异。 6.R57、R59和BobW址加的组织培养特性有一定的差异。在相同培养条件下，Bob认七i仍成熟胚的成愈率达1 00%，愈伤组织分化率为94.63%，和R57相比，两者存在显着差异。但和R59相比不存在显著差异。对去胚芽成熟胚而言，Bobwhi忆的成愈率为100%，分化率为21 .6%，R59的成愈率为91.93%，分化率为n.67%，在愈伤组织形成能力方面两者不存在显著差异，但Bobw坛妞的愈伤组织分化能力比R59强，且两者存在显著差异。 7.综上研究结果表明，R57、R59成熟胚不仅具有很高的成愈率和分化率。而且两个品种的农艺性状优良。因此，这两个小麦新品种是小麦转基因的良好受体。本研究建立的小麦商效再生体系可以为其他研究者提供参考。更多还原

【Abstract】 Mature wheat embryos of new wheat varieties R57(ChuanNongl7) and R59(ChuanNongl2) which are important cultivar in Sichuan Province were used as explants for culture in vitro.And the mature embryos of other wheat genotype"Bobwhite" were also used to culture. When mature embryos were inoculated ,some embryonic buds of RS9 and Bobwhite’ s mature embryos were removed.The purpose of this study is to construct a high effective system of plantlet regeneration for mature wheat embryos.In this study,the response of RS7 and RS9 to culture was compared with the response of Bobwhite to culture.The general result is following:In this study,a high effective wheat plantlet regeneration system was built. This experiment showed that the callus induction medium supplemented with 2mg/l 2,4-D,callus induction time of 14 days and regeneration medium with Smg/1 KT were the best combination and the best explants were mature embryos with embryonic buds.R57 and R59 have a good response to culture.The rate of callus formation of R57’ s mature embryos is 77.28%~97.58%,as for R59,the rate is 79.22%-99.33%.The highest frequency of plantlet regeneration of callus initiated from R57’ s mature embryos is 73.63%,as for R59,the highest frequency is 89.23%.Few researchers got the same result in previous studies.Harvesting time and treating by low temperature had effect on callus formation.The rate of callus formation of mature embryos excised from seeds harvested hi 2002 was higher than that of mature embryos excised from seeds harvested in 2001 .There was no significant difference in different callus induction media for callus formation of mature embryos harvested hi 2001.But great difference hi different callus induction media wasobserved for callus formation of mature embryos harvested in 2002.During the callus induction, the germination of mature embryos was affected by harvesting time and low temperature treat. It is the first time to report this kind of result The rate of germination of mature embryos harvested hi 2002 was by far higher than that of mature embryos harvested in 2001. In this study,high density of 2,4-D could prevent mature embryos from germinating,but KT could prompt germination.Embryonic bud profited callus formation and differentiation.For R59,significant difference between mature embryos with embryonic buds and those without embryonic buds was observed for callus initiation and differentiation.For Bobwhite,there was no difference between the two kinds of mature embryos for callus formation,but great diference was observed for plantlet regeneration.There were no reports about the effect of embryonic bud on callus initiation and plantlet regneration in previous studies.There was great difference between RS7 and Bobwhite for callus initiation and plantlet regeneration from mature embryos with embryonic bud.Bobwhite was better than R57.But for the same case.there was no great difference between R59 and Bobwhite.So far as mature embryos without embryonic buds was concerned,there was no significant difference between R59 and Bobwhite for callus formation,but great dufference was observed for plantlet regeneration and Bobwhite was better than R59.The new wheat varieties R57 and R59 have an excellent character for culture in vitro.And they have good agricultural trait.So they are good acceptors for transgenic wheatThe method of mature wheat embryos culture in vitro found in this study can be reference for other researchers.更多还原