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作物学报

作物学报 ›› 2022, Vol. 48 ›› Issue (10): 2435-2442.doi: 10.3724/SP.J.1006.2022.11088

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

HvLBD19基因对大麦不定根发育的调控

郭宝健1,2(), 王爽1,2, 吕超1,2, 王菲菲1,2, 许如根1,2,*()   

  1. 1江苏省作物基因组学和分子育种重点实验室 / 植物功能基因组学教育部重点实验室 / 江苏省作物遗传生理重点实验室 江苏扬州 225009
    2扬州大学农学院 / 江苏省粮食作物现代产业技术协同创新中心, 江苏扬州 225009
  • 收稿日期:2021-10-12 接受日期:2022-02-25 出版日期:2022-10-12 网络出版日期:2022-03-22
  • 通讯作者: 许如根
  • 作者简介:第一作者联系方式: E-mail: bjguo@yzu.edu.cn
  • 基金资助:
    国家自然科学基金项目(31771771);江苏省高等学校自然科学研究重大项目(19KJA560005);财政部和农业农村部国家现代农业产业技术体系建设专项(CARS-05);江苏高校优势学科建设工程项目资助

Regulation of adventitious root development by HvLBD19 gene in barley (Hordeum vulgare L.)

GUO Bao-Jian1,2(), WANG Shuang1,2, LYU Chao1,2, WANG Fei-Fei1,2, XU Ru-Gen1,2,*()   

  1. 1Jiangsu Key Laboratory of Crop Genomics and Molecular Breeding / Key Laboratory of Plant Functional Genomics of the Ministry of Education / Jiangsu Key Laboratory of Crop Genetics and Physiology, Agricultural College of Yangzhou University, Yangzhou 225009, Jiangsu, China
    2Jiangsu Co-Innovation Center for Modern Production Technology of Grain Crops, Yangzhou University, Yangzhou 225009, Jiangsu, China
  • Received:2021-10-12 Accepted:2022-02-25 Published:2022-10-12 Published online:2022-03-22
  • Contact: XU Ru-Gen
  • Supported by:
    National Natural Science Foundation of China(31771771);Natural Science Foundation of the Jiangsu Higher Education Institutions of China(19KJA560005);China Agriculture Research System of MOF and MARA(CARS-05);and the Priority Academic Program Development of Jiangsu Higher Education Institutions

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摘要：

植物LBD基因家族是植物特异的转录因子家族, 在调控植物生长发育和氮素代谢方面起到重要的作用。基因组共线性分析表明, 大麦(Hordeum vulgare L.) HvLBD19基因为水稻ARL1和玉米RTCS基因直系同源基因。基因时空表达分析结果表明, 该基因在不定根中表达丰度最高, 且受外源生长素诱导表达, 所编码蛋白定位于细胞核内。转基因功能验证结果表明, 苗期超表达株系相对于野生型不定根长度增长近1倍, 不定根数目增加40%。本研究初步明确了HvLBD19基因影响大麦不定根发育, 为进一步深入探究大麦HvLBD19基因调控不定根发育的分子机制提供了基础。

关键词: 大麦, 侧生器官边界域基因, 超表达, 不定根

Abstract:

LBD gene family is a plant specific transcription factor family, which plays an important role in regulating plant development and nitrogen metabolism. Micro-synteny analysis showed that barley HvLBD19 was an orthologous of maize RTCS and rice ARL1 gene. Temporal and spatial expression analysis revealed that this gene was most abundant in adventitious roots and was induced by exogenous auxin treatment, which encoded a protein located in the nucleus. Transgenic results indicated that the adventitious root length increased by nearly one time, and the number of adventitious roots increased by 40%. It was preliminarily confirmed that HvLBD19 gene controlled adventitious root development in barley and provided a foundation for further research on the molecular mechanism of HvLBD19 gene in regulating adventitious root development in barley.

Key words: barley, LATERAL ORGAN BOUNDARIES DOMAIN gene, overexpression, adventitious root

表1

本实验中用到的引物"

引物编号
Primer pairs ID		 引物序列
Primer sequence (5′-3′)		 内容
Content
P1 ATGACGGGACTGGGTTCGCC 编码区扩增
Coding sequence amplification
TTACGAGCGATTAAGGTAAG
P2 AGTTTCTGCGTCGCAAGTG 荧光定量
qRT-PCR
GAGCTTGGAGACGTTGCTG
P3 GGTCCATCCTAGCCTCACTC 内参基因HvActin扩增
Internal control of HvActin amplification
GATAACAGCAGTGGAGCGCT
P4 TCTCCCTTACCCATGATGAGCATGACGGGACTGGG 瞬时表达载体构建
Transient expression vector construction
CACGGGGGACTCTAGCGAGCGATTAAGGTAAGCATACGC
P5 CCCAAGCTTCTTGCATGCCTGCAGTGCAGCGTGACCCG Ubi启动子扩增
Ubi promoter amplification
CCCCCCGGGCTGCAGAAGTAACACCAAAC
P6 GACTAGTGAGCTCGAATTTCCCCGATC nos终止子扩增
nos terminator amplification
GCTCTAGAGATCTGATATCATCGATGAA
P7 CCCCCCGGGATGACGGGACTGGGTTCGCC 大麦超表达载体构建
Overexpression vector construction in barley
GACTAGTTTACGAGCGATTAAGGTAAG

图1

大麦HvLBD19基因过表达载体构建 A: 烟草瞬时表达载体构建; B: 大麦超表达载体构建。"

图2

大麦遗传转化程序 A: 幼穗; B: 幼胚愈伤诱导; C: 愈伤转化D: 幼苗再生。"

图3

LBD基因序列分析 A: HvLBD19、OsARL1和ZmRTCS蛋白序列比对; B: 玉米、大麦和水稻LBD基因的基因组区微共线性分析。"

图4

HvLBD19基因的表达分析 A: 苗期叶片(L)、不定根(AR)、根节(NR)和种子根(SR)中基因表达分析(表达量的高低以相度于叶片的表达量表示); B和C分别为NAA处理后不同时间点种子根和根节中基因表达分析(表达量以相度于0 h表达量表示)。"

图5

HvLBD19蛋白亚细胞定位 标尺为20 μm。"

图6

大麦遗传转化与阳性苗鉴定 A: 转基因株系PCR检测; P: 阳性质粒; G: Golden Promise; 数字: 不同的转基因株系; M: DNA分子量标准。B: 转基因株系潮霉素筛选。C: 阳性株系HvLBD19基因表达量分析, *和**表示转基因株系相对于野生型在0.01和0.05水平差异显著。"

图7

三叶一心期根系性状考察 A: 野生型和转基因株系苗期表型; B: 种子根长度; C: 种子根数目; D: 不定根长度; E: 不定根数目。"

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