展颖野生稻株高性状的遗传与<i>SD1</i>基因的等位变异分析

doi:10.7668/hbnxb.20194324

摘要/Abstract

摘要：

为了探究SSSL-B50和华粳籼74(HJX74)株高差异的原因,利用SSSL-B50与HJX74回交构建F₂群体进行株高性状遗传分析与基因定位。结果显示,F₁植株表现为高秆;260个单株组成的F₂群体株高出现分离,高秆植株与半矮秆植株分离比例符合3:1(χ²=0.18<3.84),说明SSSL-B50高秆性状为显性性状,受1对显性等位基因控制。利用IciMapping软件对F₂进行连锁分析,将SSSL-B50携带的高秆基因定位在1号染色体代换片段上的标记S18和X161之间38.38~39.07 Mb。候选基因筛选分析发现,定位区间内存在着“绿色革命”基因座位SD1。对展颖野生稻、SSSL-B50及HJX74进行SD1基因测序及序列比对,发现HJX74的CDS序列与展颖野生稻、SSSL-B50相比有280 bp的缺失,导致氨基酸编码提前终止。qRT-PCR结果表明,SD1表达量在SSSL-B50茎秆的第2,3,4节均显著高于HJX74。此外,与前人报道结果相比,展颖野生稻SD1基因编码的氨基酸序列发生了2处改变(E100G、Q339R)。研究结果弄清楚了展颖野生稻单片段代换系SSSL-B50高秆性状受SD1调控,同时从展颖野生稻鉴定到了SD1的新等位型SD1^Glu。

关键词: 展颖野生稻, 株高, 染色体片段代换系, 基因定位, 等位型

Abstract:

In order to investigate the reasons for the differences in plant height between SSSL-B50 and HJX74,an F₂ population was constructed by backcrossing SSSL-B50 with HJX74 for genetic analysis and gene mapping of the plant.The genetic analysis showed that F₁ plants exhibited tall plant height.And the separation ratio within F₂ was 3:1(χ²=0.18<3.84)between the tall plant individuals and the semi-dwarf ones,which indicated that the tall plant height of SSSL-B50 was a dominant trait.Based on the linkage analysis between the marker genotypes and phenotypes of plant height in the F₂ population,the gene controlling tall plant height was mapped between markers S18 and X161 within the 38.38—39.07 Mb interval of the substituted segment on chromosome 1.Moreover,it was found that the known gene of "Green revolution",SD1,fell to the mapping interval.Through amplification and sequence alignment of SD1 gene in Oryza glumaepatula,SSSL-B50,and HJX74,it showed that a 280 bp deletion in CDS in HJX74 compared with Oryza glumaepatula and SSSL-B50.The qRT-PCR results showed that the expression level of SD1 was significantly higher in the second,third,and fourth sections of SSSL-B50 stem than in HJX74.The comparison of the SD1 gene sequence of Oryza glumaepatula with the previous reported results showed that the base substitutions occurred at two sites (E100G, Q339R) in the encoded amino acid.The results revealed that the tall plant height of SSSL-B50 was controlled by SD1.Furthermore,a new allelic type of SD1, SD1^Glu,was identified in Oryza glumaepatula.

Key words: Oryza glumaepatula, Plant height, Single segment substitution line, Gene mapping, Allelic type

杨先冬, 刘伟, 曹利霞, 李孝辉, 陆怡, 傅雪琳. 展颖野生稻株高性状的遗传与SD1基因的等位变异分析[J]. 华北农学报, 2024, 39(1): 18-26. doi: 10.7668/hbnxb.20194324.

YANG Xiandong, LIU Wei, CAO Lixia, LI Xiaohui, LU Yi, FU Xuelin. Genetic Analysis of Plant Height and Allelic Variation of SD1 in Oryza glumaepatula[J]. Acta Agriculturae Boreali-Sinica, 2024, 39(1): 18-26. doi: 10.7668/hbnxb.20194324.

http://www.hbnxb.net/CN/Y2024/V39/I1/18

图/表 8

表1 本研究所用的引物

Tab.1 Primers used in this experiment

引物名称 Primer name	引物序列(5'—3') Primer sequence(5'—3')	用途 Purpose
RM486-F	CCCCCCTCTCTCTCTCTCTC	定位
RM486-R	TAGCCACATCAACAGCTTGC	定位
S18-F	CTAAATGAGACTAATTTGG	定位
S18-R	CTGTTCCAATCACAAACAATGG	定位
X161-F	GTATTGAATTGTTTCGGG	定位
X161-R	GAATTGCGATGAAGAGATAG	定位
X197-F	GTCCAAACTCTTCCATGAAG	定位
X197-R	GATCGACCATGACTGCTCT	定位
PSM369-F	GAGCCACTGATGCACCGAA	定位
PSM369-R	CGCCACCTTCACCTTCTTGT	定位
S1-F	GCGTCACGTAATGGATGGAGTATTATACAACACC	克隆SD1基因
S1-R	GTTAGGAGTAGCTCTGATGTAGATAAGCC	克隆SD1基因
SD2-F	TGTGCGACCTGAGGATGGAG	qRT-PCR
SD2-R	ACGTCGACCACGGGCATGTCCAG	qRT-PCR

图1 各供试材料的株型及株高比较 B. HJX74、SSSL-B50和展颖野生稻均为2020—2022年多季节的平均株高;F₁是2020年晚季10个单株的株高平均值。不同大写字母代表差异极显著(P<0.001)。图2,6,7同。

Fig.1 Comparison of plant types and plant heights among the tested materials B. Comparision of the average plant height among the materials in 2020—2022; in which the plant height value of F₁ was the average of 10 individual plants in the late season of 2020.Different capital letters represented the significant differences at P<0.001. The same as Fig.2,6,7.

图2 F₂群体的株高分布 A.F₂群体的株高频次分布图;B.F₂群体中3种基因型的单株数;C.F₂群体中各基因型单株的平均株高。

Fig.2 The distribution of plant height in F₂ population A.Distribution frequency of the plant height in F₂ population; B. Number of the individual plants in different genotype; C. Average plant height of three genotypes in F₂.

图3 SSSL-B50高秆基因的定位图示

Fig.3 The linkage map of the gene for tall plant height in SSSL-B50

图4 HJX74和展颖野生稻SD1基因结构变异及氨基酸序列比对 A. HJX74和展颖野生稻SD1基因结构变异图示:黑色填充框. 外显子;白色框. 内含子;灰色填充. 5'UTR和3'UTR;△和▽.HJX74与展颖野生稻相比有碱基缺失和插入。B.SD1编码的氨基酸序列比对。

Fig.4 Variations of SD1 in HJX74 and Oryza glumaepatula(Glu) and the alignment of the encoded amino acid sequences A. Variations of SD1 in HJX74 and Oryza glumaepatula gene:the black filled boxes. Exons;White boxes. Introns;The gray filled boxes. 5'UTR and 3'UTR;△ and ▽. Base deletions and insertions in HJX74 compared with Oryza glumaepatula, respectively.B.Alignment of the amino acid sequences encoded by SD1 between HJX74 and Oryza glumaepatula.

图5 不同水稻材料SD1基因编码的氨基酸序列差异 NIP、TC65和Glu分别代表Nipponhare、Taichung 65和展颖野生稻。

Fig.5 Differences in amino acid sequences encoded by SD1 genes in different rice germplasms NIP, TC65, and Glu represented Nipponhare, Taichung 65, and Oryza glumaepatula, respectively.

图6 HJX74和SSSL-B50节间长比较及SD1基因在各茎节的相对表达量 A.HJX74和SSSL-B50节间长比较图;B,C.HJX74和SSSL-B50各节间在不同生育时期的节间长;Ⅰ—Ⅵ分别代表水稻顶部的第1节到第6节;D.孕穗时期SD1基因在HJX74、SSSL-B50各节间的相对表达量。不同小写字母代表差异显著(P<0.05)。

Fig.6 Comparison of the internodal lengths between HJX74 and SSSL-B50,and the relative expression levels of SD1 in the internodes A.HJX74 and SSSL-B50 internode elongation pattern diagram; B, C.The internode length of HJX74 and SSSL-B50 at different growth stages; Ⅰ—Ⅵ indicated the first to the sixth internode; D. Relative expression levels of SD1 in the internodes of HJX74 and SSSL-B50 during booting stage. Different lowercase letters represented the significant differences at P<0.05.

图7 HJX74在外施不同浓度GA₃条件下的株高变化

Fig.7 Plant height of HJX74 under the condition of external GA₃ application

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展颖野生稻株高性状的遗传与SD1基因的等位变异分析

Genetic Analysis of Plant Height and Allelic Variation of SD1 in Oryza glumaepatula

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