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作物学报 ›› 2020, Vol. 46 ›› Issue (11): 1743-1749.doi: 10.3724/SP.J.1006.2020.01016

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

小麦高效转基因受体品系CB037的抗条锈性分析

郑燕燕(), 黄德华, 李金龙, 张会飞, 鲍印广, 倪飞, 吴佳洁*()   

  1. 山东农业大学农学院 / 作物生物学国家重点实验室, 山东泰安 271018
  • 收稿日期:2020-02-24 接受日期:2020-06-02 出版日期:2020-11-12 网络出版日期:2020-06-15
  • 通讯作者: 吴佳洁
  • 作者简介:E-mail:zyy18854889722@163.com, Tel: 0538-8242279
  • 基金资助:
    本研究由国家转基因生物新品种培育重大专项(2016ZX08009003001);山东省高等学校“青创科技计划”项目(2019KJF026)

Analysis of the stripe rust resistance in a wheat line CB037 with high regeneration and transformation efficiency

ZHENG Yan-Yan(), HUANG De-Hua, LI Ji-Long, ZHANG Hui-Fei, BAO Yin-Guang, NI Fei, WU Jia-Jie*()   

  1. State Key Laboratory of Crop Biology / College of Agronomy, Shandong Agricultural University, Tai'an 271018, Shandong, China
    State Key Laboratory of Crop Biology / College of Agronomy, Shandong Agricultural University, Tai’an 271018, Shandong, China
  • Received:2020-02-24 Accepted:2020-06-02 Published:2020-11-12 Published online:2020-06-15
  • Contact: Jia-Jie WU
  • Supported by:
    This study was supported by the National Major Project for Developing New GM Crops(2016ZX08009003001);the Youth Innovation and Scientific Program of Higher Education of Shandong(2019KJF026)

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摘要：

小麦遗传转化是开展小麦基因克隆、基因编辑及基因工程等研究的重要基础。获得易于转化及遗传背景纯合的受体材料是高效开展小麦遗传转化的关键。近年来, 小麦品系CB037作为受体材料被广泛利用, 并获得了较高的转化效率, 在小麦研究中发挥了重要作用。虽然野生型CB037农艺性状整齐一致, 但遗传背景并不纯合, 对小麦条锈菌(Puccinia striiformis f. sp. tritici)表现高感和高抗两种不同反应。本研究分离了抗病株系CB037-PstR及感病株系CB037-PstS, 并创建了F2分离群体。遗传分析表明, CB037-PstR携带单个显性抗病基因。利用BSR-Seq测序分析及分子标记检测, 将抗病位点定位于1B染色体短臂。进一步荧光原位杂交表明, CB037-PstR为1BL/1RS易位系, 可能携带抗条锈病基因Yr9。本研究明确了CB037的遗传背景及其抗条锈性, 并分离了抗病及感病株系, 为有效利用CB037作为受体材料开展小麦转基因研究奠定基础。

关键词: 小麦遗传转化, 1BL/1RS易位系, 条锈病抗性, BSR-Seq, Yr9

Abstract:

Genetic transformation is used for efficient gene cloning, gene editing and gene engineering, etc. Obtaining recipient lines amenable to transformation and with pure genetic background is critical for high efficiency transformations. For recent years, the wheat inbreed line CB037 had been widely used as a recipient for transgenes and obtain its high transformation potential. Despite having stable agronomic traits, the CB037 is genetically heterogeneous for resistance to wheat stripe rust (Puccinia striiformis f. sp. tritici, Pst). In this study, the Pst-resistant line CB037-PstR and Pst-susceptible line CB037-PstS were isolated, and their F2 population was created. Genetic analysis showed that the CB037-PstR carried a single dominant resistance gene. The identified resistance gene was mapped on the short arm of chromosome 1B using BSR-seq and molecular marker analysis. GISH results further revealed that CB037-PstR is a 1BL/1RS translocation line and likely carried Yr9. This study segregated genetic heterogeneity in CB037 for stripe rust resistance and isolated its Pst-susceptible and resistant lines. These results will facilitate trait-oriented use of CB037 in genetic engineering of wheat.

Key words: wheat transformation, 1BL/1RS translocation line, stripe rust resistance, BSR-Seq, Yr9

图1

CB037所分离出的两个株系对条锈菌的不同反应 1, 2: 苗期叶片; 3: 成株期叶片。"

图2

抗病株系CB037-PstR与感病株系CB037-PstS及F2感病单株混合池间差异位点的染色体分布图 竖线及绿色圆点分别代表小麦染色体及着丝粒。上方数字及字母代表染色体编号, 下方数字为定位于该染色体的SNP及InDel数量, 短横线代表SNP及InDel所在位置。左侧标尺代表染色体长度, 单位Mb。"

表1

本研究开发的分子标记"

分子标记
Molecular marker		 染色体及位置
Chromosome & position (Mb)		 正向引物
Forward primer
(5′-3′)		 反向引物
Reverse primer
(5′-3′)		 退火温度
Annealing temperature (℃)		 内切酶
Restriction enzyme		 多态性
Polymorphism
(bp)
CBSR-53/54 a 1B, 69.9 GAACGGCTCGGATCTGATC TGTACAACAACAAGAAGGAGGAG 55 Dde I CBS: 79+83
CBR: 163
CBSR-1/2 b 1B, 98.7 CCGGAGTTGAAGATAAGTAGTTG CTTCCCTGTATTCGAACAGACTCA 56 N/A CBS: 242
CBR: 0
CBSR-35/36 a 1B, 115.3 ACACCGAGGTGCGTCTAAT TATCCCATGCGGTAAATGGAC 55 Msp I CBS: 150+57
CBR: 207
InDel-3/4 b 1B, 376.6 TCCATGGCAGTACCGCTTGCTT CCGGAGCAGCAATGAAACGTC 56 N/A CBS: 240
CBR: 207
CBSR-25/26 b 4A, 723.7 GCCGCGTTTTGCATGTTTTCCG TGGGTTGAATTTTGGGTTGTTTGAA 56 N/A CBS: 408
CBR: 378

图3

抗条锈病基因Yr9分子标记(iag95, AF1/4, H20)的检测结果 S、R分别表示F2个体的感病及抗病表型; CK为空白对照。"

图4

CB037两个株系的染色体荧光原位杂交鉴定 A, B: CB037-PstS; C, D: CB037-PstR; A和C: GISH鉴定; B和D: FISH鉴定。标尺长度为10 μm。"

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