大豆成熟种子萌发叶片体细胞胚的高频诱导和遗传转化
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1.1)青岛农业大学生命科学学院,山东省高校植物生物技术重点实验室,青岛 266109;2.2)山东宝来利来生物工程股份有限公司,泰安 271000

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Q7

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山东省自然科学基金(ZR2019MC033)资助项目。


High Frequency Induction and Transformation of Somatic Embryos From Mature Seed-derived Leaf in Soybean
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1.1)College of Life Sciences, Key Lab of Plant Biotechnology in Universities of Shandong Province, Qingdao Agricultural University, Qingdao 266109, China;2.2)Shandong Baolai-Leelai Bio-Industrial Group, Taian 271000, China

Fund Project:

This work was supported by a grant from Natural Science Foundation of Shandong Province (ZR2019MC033).

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    摘要:

    目的 虽然大豆遗传转化取得了较大进展,但其转化效率仍然偏低。因此,有必要建立一个更加简单和高效的大豆转化系统。本研究的目的是以大豆成熟种子萌发产生的叶片(MSDL)为靶外植体,高频诱导和转化大豆体细胞胚(somatic embryos,SEs)。方法 萌发7 d的大豆幼苗上摘取叶片,并将其切成1.2 cm×1.2 cm的外植体。将叶片外植体置于胚胎诱导培养基(EIM)上诱导体细胞胚发生。将叶片外植体浸入携带pCAMBIA1301植物表达载体的农杆菌EHA105菌液进行遗传转化。结果 MSDL(包括初生叶和次生叶)均能被诱导产生SEs。大多数SE发生在叶片切口处。所试的不同基因型均能诱导SEs。SEs的最高诱导率为95.0%。β葡糖醛酸糖苷酶(GUS)染色结果表明,平均转化效率达到75.4%。DNA印迹(Southern blot)结果表明,目的基因稳定整合在大豆基因组中,拷贝数为1~3个。结论 本研究建立了以大豆MSDL为靶组织的体细胞胚高频诱导和遗传转化。该系统有望在大豆基因组编辑技术的开发上得到应用。

    Abstract:

    Objective Although many improvements in soybean transformation have been made, the efficiency of transformation is still low. Thus, a more useful and efficient soybean transformation system required to be established. The objective of this study is to develop high frequency induction and transformation of somatic embryos (SEs) using mature seed-derived leaf (MSDL) of soybean as target explants.Methods The leaves detached from 7-d-old germinated seedlings were divided into 1.2 cm×1.2 cm sections. These sections were used to induce somatic embryogenesis on the embryo induction medium (EIM) and to infect with Agrobacterium EHA105 strain harboring the pCAMBIA1301 for transformation.Results The MSDL including primary leaf and secondary leaf was able to produce SEs. Most of SEs occurred along the incision of leaves. This system was shown to be applicable to different genotypes. The highest induction frequency of SEs was 95.0% and the average transformation efficiency based on β-glucuronidase (GUS) assay reached to 75.4%. The transgenic events were further con?rmed by Southern blot.Conclusion High frequency induction and transformation of somatic embryos using soybean MSDL as target tissue were developed in this study. This system can be applied to the development of genome editing technology in soybean.

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孙明杰,侯恒,赵春梅,薛仁镐.大豆成熟种子萌发叶片体细胞胚的高频诱导和遗传转化[J].生物化学与生物物理进展,2023,50(8):1937-1945

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  • 收稿日期:2022-11-14
  • 最后修改日期:2023-01-30
  • 接受日期:2023-01-30
  • 在线发布日期: 2023-08-14
  • 出版日期: 2023-08-20