不依赖基因型的高效玉米遗传转化体系的建立

doi:10.3724/SP.J.1006.2022.13068

作物学报 ›› 2022, Vol. 48 ›› Issue (12): 2987-2993.doi: 10.3724/SP.J.1006.2022.13068

• 作物遗传育种·种质资源·分子遗传学 • 上一篇下一篇

不依赖基因型的高效玉米遗传转化体系的建立

许洁婷¹^,²^,³(), 刘相国⁴(), 金敏亮¹^,², 潘弘², 韩宝柱², 李梦娇², 岩说⁵, 胡国庆⁵, 严建兵¹()

¹华中农业大学作物遗传改良国家重点实验室, 湖北武汉 430070
²未米生物科技(江苏)有限公司, 江苏常州 213000
³未米生物科技(海南)有限公司, 海南三亚 572000
⁴吉林省农业科学院农业生物技术研究所, 吉林长春 130033
⁵西双版纳傣族自治州农业科学研究所, 云南景洪 666100

收稿日期:2021-11-26 接受日期:2022-03-25 出版日期:2022-12-12 网络出版日期:2022-04-20
通讯作者: 严建兵
作者简介:许洁婷, E-mail: xjt@wimibio.com;
刘相国, E-mail: lxgyyj@cjaas.com第一联系人：
**同等贡献
基金资助:
国家转基因生物新品种培育重大专项(2018ZX08010-04B);国家转基因生物新品种培育重大专项(2019ZX08010003-002-013);吉林省科技发展计划项目(20190201290JC)

Establishment of genotype-independent high-efficiency transformation system in maize

XU Jie-Ting¹^,²^,³(), LIU Xiang-Guo⁴(), JIN Min-Liang¹^,², PAN Hong², HAN Bao-Zhu², LI Meng-Jiao², YAN Shuo⁵, HU Guo-Qing⁵, YAN Jian-Bing¹()

¹National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan 430070, Hubei, China
²WIMI Biotechnology Co., Ltd., Changzhou 213000, Jiangsu, China
³WIMI Biotechnology Co., Ltd., Sanya 572000, Hainan, China
⁴Institute of Agricultural Biotechnology, Jilin Academy of Agricultural Sciences, Changchun 130033, Jilin, China
⁵Institute of Agricultural Sciences of Xishuangbanna Prefecture of Yunnan Province, Jinghong 666100, Yunnan, China

Received:2021-11-26 Accepted:2022-03-25 Published:2022-12-12 Published online:2022-04-20
Contact: YAN Jian-Bing
About author:First author contact:
**Contributed equally to this work
Supported by:
National Major Project for Developing New GM Crops(2018ZX08010-04B);National Major Project for Developing New GM Crops(2019ZX08010003-002-013);Science and Technology Innovation Program of Jilin Province(20190201290JC)

摘要/Abstract

摘要：

玉米遗传转化严重依赖于受体基因型, 目前绝大部分商业化玉米骨干自交系的遗传转化仍然十分困难。Baby boom (Bbm)和Wuschel2 (Wus2)是植物干细胞发育中的2个关键基因, 调节这2个基因的时空表达可以有效提高植物遗传转化效率。本研究以多个我国玉米骨干自交系为受体材料开展转化效率测试, 发现虽然Bbm和Wus2的超表达可以显著提高转化效率, 但对玉米的生长发育具有不利影响。本研究开发了一种新型辅助转化技术, 在辅助载体中加入致死基因表达元件, 并调节2个基因的时空表达。结果表明该辅助载体和目标载体混合转化时, 不但可以在T₀代成功获得不含Bbm和Wus2辅助载体的正常生长发育的优质转化苗, 还显著提高转化效率(平均达19.5%)。本研究成功建立和完善了一套不依赖基因型的高效玉米遗传转化体系, 为提高生物育种效率和精准改良提供了有力支持。

关键词: 玉米, 遗传转化, Baby boom, Wuschel2, 辅助载体

Abstract:

The reliance on receptor genotype of genetic transformation made it difficult for the transformation of commercial maize lines. Expression regulation of two important genes in plant stem cell development, Baby boom (Bbm) and Wuschel2 (Wus2), was revealed to significantly improve transformation efficiency. Several Chinese core maize inbred lines were used as receptor materials to test the transformation efficiency. Although the overexpression of Bbm and Wus2 could significantly improve the transformation efficiency, it had a negative impact on the growth and development of T₀ plants. Here, a new assisted transformation technology was developed, in which a lethal gene element was added to the assist vector and the spatial expression of the two genes were regulated. The results revealed that the hybrid transformation of the assist vector and the target vector could not only successfully obtain high-quality transformation seedlings without Bbm and Wus2 assist vector in T₀ generation, but also significantly improve the transformation efficiency with an average 19.5%. The application of this improved genotype-independent genetic transformation system promises maize precise improvement with higher efficiency.

Key words: maize, genetic transformation, Baby boom, Wuschel2, assist vector

许洁婷, 刘相国, 金敏亮, 潘弘, 韩宝柱, 李梦娇, 岩说, 胡国庆, 严建兵. 不依赖基因型的高效玉米遗传转化体系的建立[J]. 作物学报, 2022, 48(12): 2987-2993.

XU Jie-Ting, LIU Xiang-Guo, JIN Min-Liang, PAN Hong, HAN Bao-Zhu, LI Meng-Jiao, YAN Shuo, HU Guo-Qing, YAN Jian-Bing. Establishment of genotype-independent high-efficiency transformation system in maize[J]. Acta Agronomica Sinica, 2022, 48(12): 2987-2993.

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试验编号 Test ID	玉米自交系 Maize inbred line	载体 Vector name	起始胚数量 No. of embryos	转化阳性苗数量 No. of positive transgenic seedlings	转化效率 Transformation frequency (%)
1	郑58 Zheng 58	pWMDR001	100	24	24.0
2	X923-1	pWMDR001	50	3	6.0
3	IL3	pWMDR001	50	5	10.0
4	IL4	pWMDR001	50	9	18.0
5	京724 Jing 724	pWMDR001	50	6	12.0
6	KN5585	pWMDR001	50	9	18.0
7	B104	pWMDR001	100	18	18.0
8	郑58 Zheng 58	p193412	200	0	0
9	X923-1	p193412	100	0	0
10	IL3	p193412	100	0	0
11	IL4	p193412	100	0	0
12	京724 Jing 724	p193412	100	0	0
13	KN5585	p193412	200	21	10.5
14	B104	p193412	100	5	5.0

试验编号 Test ID	玉米自交系 Maize inbred line	起始胚数量 No. of embryos	转化阳性苗数量 No. of positive transgenic seedlings	优质转化苗数量 No. of high-quality transformed seedlings	优质转化苗分离率 Separation rate of high-quality transformed seedling (%)	优质转化效率 High-quality transformation frequency (%)
1	郑58 Zheng 58	313	24	8	33.3	2.6
2	郑58 Zheng 58	535	107	2	1.9	0.4
3	郑58 Zheng 58	828	129	56	43.4	6.8
4	郑58 Zheng 58	446	103	14	13.6	3.1
5	KN5585	210	40	5	12.5	2.4
6	KN5585	256	65	18	27.7	7.0
7	KN5585	260	51	16	31.4	6.2
8	B104	189	59	13	22.0	6.9
9	B104	242	70	11	15.7	4.5
10	B104	150	48	10	20.8	6.7

试验编号 Test ID	玉米自交系 Maize inbred line	起始胚数量 No. of embryos	转化阳性苗数量 No. of positive transgenic seedlings	优质转化苗数量 No. of high-quality transformed seedlings	优质转化苗分离率 Separation rate of high-quality transformed seedling (%)	优质转化效率 High-quality transformation frequency (%)	平均优质转化效率 Average high-quality transformation frequency (%)
1	B104	430	133	123	92.5	28.6	31.0
2	B104	100	39	39	100.0	39.0
3	B104	150	45	38	84.4	25.3
4	郑58 Zheng 58	160	18	14	77.8	8.8	16.4
5	郑58 Zheng 58	200	56	48	85.7	24.0	16.4
6	京724 Jing 724	50	6	5	83.3	10.0	11.5
7	京724 Jing 724	100	15	13	86.7	13.0	11.5
8	糯IN12 Nuo IN12	50	7	7	100.0	14.0	13.3
9	糯IN12 Nuo IN12	200	27	25	92.6	12.5	13.3

不依赖基因型的高效玉米遗传转化体系的建立

Establishment of genotype-independent high-efficiency transformation system in maize

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