* 通讯作者(Corresponding author): 张新友, E-mail:haasz@126.com, Tel: 0371-65729560 **同等贡献(Contributed equally to this work) 第一作者联系方式: E-mail:lilina198283@163.com, Tel: 0371-65718247 收稿日期:2016-09-18 基金:本研究由河南省重大科技专项(141100110600), 国家现代农业产业技术体系建设专项(CARS-14)和河南省现代农业产业技术体系项目(S2012-5)资助
摘要花生野生种是改良花生栽培种的重要基因资源。为了利用野生花生的抗性基因, 本研究利用花生栽培品种豫花9331与二倍体野生种 A. oteroi人工杂交, 借助胚拯救和染色体秋水仙素加倍, 创制一个双二倍体杂种AmE-4, 并利用荧光原位杂交和分子标记技术准确鉴定了该双二倍体。观察结果表明, AmE-4的叶片与豫花9331存在显著差异, 而主茎高、侧枝长和总分枝数等性状与豫花9331差异不显著。AmE-4开花期较豫花9331推迟60 d, 结实性与荚果发育状况较差, 不利于AmE-4的育种利用。同时, 开发了57个追踪AmE-4中 A. oteroi染色体的显性或共显性SSR标记, 为创制和鉴定花生栽培种 A. oteroi易位系或渐渗系奠定分子基础。
关键词:花生; 双二倍体; 花生野生种; 分子标记; 荧光原位杂交 Development and Characterization of Amphidiploid Derived from Interspecific Cross between Cultivated Peanut and Its Wild Relative Arachis oteroi LI Li-Na1,2,**, DU Pei2,**, FU Liu-Yang2,3, LIU Hua2, XU Jing2, QIN Li2, YAN Mei2, HAN Suo-Yi2, HUANG Bing-Yan2, DONG Wen-Zhao2, TANG Feng-Shou2, ZHANG Xin-You2,* 1College of Agricultural, Henan University of Science and Technology, Luoyang 471023, China
2 Industrial Crops Research Institute, Henan Academy of Agricultural Sciences / Key Laboratory of Oil Crops in Huanghuaihai Plains, Ministry of Agriculture / Henan Provincial Key Laboratory for Oil Crops Improvement, Zhengzhou 450002, China
3 School of Life Sciences, Zhengzhou University, Zhengzhou 450001, China
Fund:This study was supported by the Major Technology Research and Development Program of Henan Province (141100110600), the China Agriculture Research System (CARS-14), and the Henan Provincial Agriculture Research System (S2012-05) AbstractWild Arachis species are important genetic resources. To introgress resistant genes of Arachis species, we developed a new amphidiploid AmE-4 through man-made cross between cultivated peanut variety Yuhua 9331 and a diploid Arachis species A. oteroi, with the assistance of following embryo rescue and chromosome doubling by colchicine treatment. AmE-4 was identified and characterized by fluorescence in situ hybridization (FISH) and SSR molecular marker. Morphological observation revealed significant differences in leaves between amphidiploid AmE-4 and Yuhua 9331, while the agronomic traits such as main stem height, length of first lateral branch and number of branches showed less difference between them. The date of first flower appearance in AmE-4 delayed sixty days compared with that in Yuhua 9331, and its pods setting and development were also poor, which would hinder its further utilization. In addition, 57 dominant or co-dominant SSR molecular markers were developed and could be used to identify translocation or introgression lines with A. oteroi chromosome fragment in future studies.
Keyword:Peanut ( Arachis hypogaea L.); Amphidiploid; Wild Arachis species; Molecular marker; FISH Show Figures Show Figures
图1 S0有果针枝(A)和无果针枝(B)花的花粉染色 黄色箭头所示为可育花粉粒, 蓝色箭头所示为不育花粉粒。Fig. 1 Pollen staining from flowers in branches with (A) and without pegs (B) Yellow arrows show fertile pollens and blue arrows show sterile pollens.
图3 豫花9331、A. oteroi和AmE-4荧光原位杂交 A~B: 豫花9331 DAPI染色及45S rDNA (红)和5S rDNA (绿)探针荧光原位杂交; C~D: A. oteroi DAPI染色及45S rDNA (红)和5S rDNA (绿)探针荧光原位杂交; E~F: AmE-4 DAPI染色和A.oteroi全基因组探针原位杂交。红色箭头所示为小染色体, 白色箭头所示为“ 大随体染色体” ; F图中的白色* 表示强基因组杂交信号的染色体, F图中的红色+表示弱基因组杂交信号的染色体。Fig. 3 Fluorescence in situ hybridization of Yuhua 9331, A. oteroi and AmE-4 A-B: DAPI staining and FISH of Yuhua9331 with 45S rDNA (red) and 5S rDNA (green) probes; C-D: DAPI staining and FISH of A. oteroi with 45S rDNA (red) and 5S rDNA (green) probes; E-F: DAPI staining and GISH of AmE-4 with A. oteroi total genomic DNA probes. Red arrows show the small chromosomes, white arrows show “ the big SAT-chromosomes” ; * show chromosomes with strong signals of A. oteroi total genomic DNA probes, and + show chromosomes with weak signals of A. oteroi total genomic DNA probes in Fig. F.
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