与SP1互作的水稻穗顶部退化基因qPAA3的精细定位

【目的】水稻顶部小穗退化减少了单穗的总枝梗数和总粒数，严重影响单株产量，是水稻生产上的一个不利性状。因其遗传基础复杂，受环境影响较大，控制顶部小穗退化的相关基因克隆研究报道极少，该不利性状发生的分子机制及其遗传网络还不得而知。对顶部小穗退化基因进行精细定位，可为穗顶部基因的克隆奠定基础；开发的紧密连锁分子标记，也可以运用于分子育种实践，对这一不利性状进行早期识别和淘汰。【方法】首先对小穗突变体sp进行精细定位。用sp分别与粳稻品种ITA182和籼稻品种J160杂交构建2个遗传定位群体。为了研究不同穗退化突变体之间的关系，再以小穗突变体sp和穗顶部退化材料05261杂交，获得了农艺性状稳定的拟双突变体（表型与sp相似）。通过连续自交，纯合拟双突变体的遗传背景。再以高代的拟双突变体为非轮回亲本，穗顶部正常品种IRAT129为轮回亲本，构建含有双突变体的BC₁F₂亚群体。其中一个亚群体14C2017既表现单基因的穗退化性状分离，又出现小穗和穗顶部退化的双突变体表型，被用作顶部小穗退化基因的精细定位材料。【结果】水稻小穗性状是由1对隐性基因（sp）控制的。利用混池方法将SP(t)初步定位于第11染色体分子标记RM26281与RM7391之间；利用新开发的60对SSR分子标记，将其定位在标记sc50和sc66之间。在此区间内设计引物，最终将SP(t)定位在标记sc24和sc66之间，物理距离为54.3 kb的范围内。测序结果表明，突变体在该区间内有15.03 kb的大片段缺失，导致基因SP1的编码序列缺失。对拟双突变体的表型分析表明，sp与一个穗顶部退化基因存在互作。利用亚群体14C2017作为克隆与SP1互作基因的遗传分离群体，利用分布于全基因组的239对引物，筛选出在拟双突变体和IRAT129之间有多态的引物114对，将目标基因精细定位于第3染色体SSR标记RM6929和RM1319之间，物理距离为97.3 kb范围内，该候选基因属于早先报道的QTL--qPAA3。【结论】水稻sp的小穗性状是由基因SP1引起的缺失突变。与SP1互作的qPAA3定位于第3染色体SSR标记RM6929和RM1319之间，物理距离为97.3 kb的范围内。

Fine Mapping of Rice Panicle Apical Abortion Gene qPAA3 Interacting with SP1

【Objective】 Rice spikelet degradation reduces branch number and total grain number of single panicle, which seriously cause the yield loss per plant, is a harmful character of rice. Because of the complex geneticbasis, together frequently influenced by the environment factors,up to now, there is few report about panicle apical abortion-related gene isolation. Fine mapping of panicle apical abortion mutant is meaningful in understanding of the molecular mechanism underlying panicle apical abortion formation and in the field of molecular breeding.【Method】In the present study, SP1 was mapped using the two F₂ populations derived from the crosses between sp and ITA182, sp and J160, respectively. qPAA3 was finely mapped using the BC₁F₂ sub-population numbered 14C2017 with a pseudo-double-mutant as the non-recurrent parent originated from the cross between sp and a panicle apical abortion line 05261.【Result】 Genetic analysis showed that the sp was determinated by a single recessive nuclear gene. The candidate gene SP(t)was primarily linked on Chr.11 with the SSR markers of RM26281 and RM7391. Using the newly developed 60 markers, the SP(t) gene was further mapped between sc50 and sc66, with the genetic distance of 0.29 cM and 0.23 cM, respectively, and the physical distance 229 kb. Finally, the SP(t)gene was finely mapped between sc24 and sc66 with the physical distance of 54.3 kb. The sequencing results showed that there was 15.03 kb deletion within this region in mutant. The candidate gene was identified as a well-known gene SP1. Then, through analysis on the phenotype of double mutants combining bt and sp, it was found that there is an interaction between one of panicle apical abortion QTLs and SP1. Over consecutive 11 generation of self-crossing, the pseudo-double-mutant was further used as recurrent backcross parent to construct 81 BC₁F₂ sub-populations. Linkage analysis on 184 individuals from the sub-population 14C2017 by 114 polymorphic SSR primers revealed that, one QTL, within the fragment of 97.3 kb delimited by the SSR markers of RM6929 and RM1319, was corresponding to the previously documented QTL-qPAA3. 【Conclusion】The small panicle mutant sp was caused by the deletion of candidate gene SP1,interacting with one of the panicle apical abortion QTLs from bt. The later was qPAA3 and finally delimited to the physical distance of 97.3 kb by SSR marker RM6929 and RM1319 on the chromosome 3.