摘要
【目的】通过对甘蓝型油菜耐盐相关性状进行全基因组关联分析,寻找可能与油菜耐盐性相关的SNP位点,发掘与油菜耐盐性有关的候选基因。【方法】以1.2%NaCl溶液作为培养液,去离子水为对照,对307个不同品系的甘蓝型油菜进行发芽试验。播种后7 d测定幼苗根长、鲜重及发芽率,计算盐胁迫下各性状相对值,并作为评价耐盐的指标。结合油菜60K SNP芯片,利用SPAGeDi v1.4软件对该群体307份甘蓝型油菜进行亲缘关系分析,并计算亲缘关系值的矩阵。利用软件STRUCTURE v2.3.4对关联群体进行了群体结构分析。为有效排除假关联的影响,将群体结构和材料间的亲缘关系考虑进关联分析中,同时进行了PCA+K模型、Q+K模型以及K模型3种混合线性模型分析和比较,根据所有SNP的–lg(P)观察值和期望值,确定每个性状GWAS分析的最优模型。采用TASSEL 5.0软件,在最优模型下对307份材料耐盐各性状的相对值分别与SNP标记进行全基因组关联分析。利用油菜基因组数据库,在显著SNP位点侧翼序列200 kb范围内提取基因。根据拟南芥中已经明确功能的耐盐相关基因,筛选出目标基因组区段内与耐盐相关的油菜同源基因。【结果】全基因组关联分析共检测到164个与根长显著关联的SNP位点,23个与鲜重显著关联的SNP位点,38个与发芽率显著关联的SNP位点。其中与根长、鲜重、发芽率最显著关联的SNP位点分别位于染色体A08、A02和A06,贡献率分别为23.84%、18.59%和31.81%。在这些显著SNP位点侧翼序列200 kb范围内发掘出可能与油菜耐盐性有关的50个候选基因。这些候选基因主要包括转录因子MYB、WRKY、ABI1、b ZIP、ERF1、CZF1、XERICO等以及一些下游受转录因子调控的不同功能基因NHX1、PTR3、CAT1、HKT、CAX1、ACER、STH、STO等。在根长和发芽率2个不同耐盐性状的分析结果中均筛选出位于A03染色体上的耐盐基因BnaA03g14410D。另外,这些耐盐候选基因中包含两组串联重复基因,分别是位于A03染色体上的BnaA03g18900D和BnaA03g18910D,位于C09染色体上的BnaC09g19080D、BnaC09g19090D和BnaC09g19100D。除此之外,耐盐候选基因中还包含2个距离非常近(中间只间隔2个基因)的重复基因BnaC02g39600D和BnaC02g39630D。【结论】共检测到225个与耐盐性状显著关联的SNP位点,筛选出50个可能与油菜耐盐性有关的候选基因。
[Objective] To find the SNP loci and candidate genes associated with salt tolerance in rapeseed, genome-wide association analysis of salt tolerance related traits in B. napus was performed. [Method] The seeds of 307 inbred lines of the association panel were germinated in Petri dishes under 1.2% NaCl. The control was under the sterile dH20. Three salt tolerances related traits, root length, fresh weight and germination rate of each line were measured seven days after planting. The relative value of root length, flesh weight and germination rate under the salt tolerance condition were calculated. The software TASSEL5.0 were employed to examine salt tolerance related traits in B. napus using a genome-wide association study with a 60K Brassica Illumina~ Infinium SNP array. The software SPAGeDi vl.4 was used to calculate the relative kinship matrix comparing all pairs of the 307 accessions. STRUCTURE v2.3.4 was employed to analyze the population structure of the association panel. In this study, to avoid the false negative associations, three mixed models controlling relative kinship, K model, Q＋K model and PCA ＋K model were chosen to determine the statistical associations between phenotypes and genotypes to evaluate the effects of population structure and relative kinship on salt tolerance related traits. The optimum model was determined according to observed P values and expected P values of all SNPs. TASSEL 5.0 was used to perform genome-wide analysis under the optimum model. Based on the physical positions of the associated SNP loci, the corresponding genomic sequences of the regions and flanking sequences （200 kb upstream and downstream of the SNPs） were extracted. The genes in the target intervals were extracted subsequently. According to the salt tolerance related genes in Arabidopsis, the homologous genes of rapeseed were screened out in the target genome interval. [Result] The association analysis identified 164 SNP loci significantly associated with the root length, 23 SNP loci with flesh weight, and 38 SNP loci with germination rate. The SNP loci which were most significantly associated with root length, flesh weight and germination rate were located on chromosomes A08, A02 and A06, and explained 23.84%, 18.59% and 31.81% of the phenotypic variance, respectively. Fifty candidate genes were found to be related to salt tolerance in the target interval of B. napus. These candidate genes include transcription factors Myb, WRKY, ABI1, bZIP, ERF1, CZF1, XERICO and some different functional genes which are regulated by the transcription factors, such as NHX1, PTR3, CAT1, HKT, CAX1, ACER, STH, STO, etc. The common salt tolerance gene BnaAO3g14410D on chromosome A03 was screened out according to two different traits, root length and germination rate. In addition, the salt tolerance candidate genes contain two pairs of tandem repeats genes which are BnaAO3g18900D and BnaAO3g18910D located on chromosome A03 and BnaCO9gl908OD, BnaCO9gl9090D and BnaCO9gl91OOD located on chromosome C09. Moreover, the salt tolerant candidate genes also contain two duplicated genes BnaCO2939600D and BnaCO2g39630D with very close distances. [Conclusion] Totally, 225 SNP loci were detected significantly associated with salt tolerance related traits and fifty candidate genes were found.
出处
《中国农业科学》
CAS
CSCD
北大核心
2017年第7期1189-1201,共13页
Scientia Agricultura Sinica
基金
国家重点研发计划"七大农作物育种"重点专项(2016YFD0100202)
国家自然科学基金(31671729
31301351)
中央高校基本科研业务费(XDJK2016B031)
