PCR标记对水稻温敏雄性核不育株的鉴别

PCR Identification of Thermo-sensitive Genic Male Sterile Plants in Rice

用3个与水稻温敏雄性核不育(TGMS)基因紧密连锁的PCR标记C365-1、S2-40和In Del37,对云南低海拔(260 m)早季种植的2个籼稻TGMS株系配制的杂交组合的F2和F3代分离群体中筛选出的146个单株(126个TGMS单株和20个可育株)及经多代自交获得的15个TGMS株系的基因型进行鉴别,结果表明:只有标记C365-1在来自两个群体的不育株和可育株DNA池间显示出多态性。不育株DNA池产生两个PCR片段,分别为418 bp(定名为C365-418 bp)和344 bp(定名为C365-344 bp),而可育株DNA池只产生C365-418 bp片段。经克隆测序比对,显示多态性的C365-344 bp片段与水稻第2染色体上的OJ1001_D05克隆的DNA序列相似性为84%,与TGMS基因ptgms2-1和tms5的距离约为19.1 kb;与水稻第6染色体上的OJ1001_B06.3和OJ1001_B06.4两个克隆间隔区的DNA序列的相似性为98%,与TGMS基因TMS相距约1 c M。在146个单株中,标记C365-1的基因型值与单株育性表型值间的相关系数为0.944,达极显著水平。来自15个TGMS株系的每一个株系都显示出与不育株DNA池相同的带型。以上结果说明,PCR标记C365-1可较为有效地鉴别水稻TGMS株携带的TGMS基因,该标记可作为水稻TGMS株基因型分子标记辅助选择的有效标记。

Three PCR markers C365-1, S2-40, and InDel37, which were tightly linked with Thermo-sensitive Genetic Male Sterile （TGMS） genes reported, were used to identify TGMS gene in 146 individuals （including 126 TGMS and 20 fertile individuals） selected from F2 and F3 genetic segregated populations, which were derived from two indica hybrid combinations made by two TGMS lines, grown in the early-season in a rice field with 260 m low altitude in Yunnan, and 15 TGMS lines through selfing-crosses for many generations. The results showed that only marker C365-1 gave polymorphism between DNA pools of sterile and fertile plants from the two segregated populations. The DNA pool of sterile plants possessed two PCR fragments, one with 418 bp （reference as C365-418 bp）, and another with 344 bp （reference as C365-344 bp）. While the fertile plant DNA pools possessed only the PCR fragment with C365-418 bp. Sequencing and alignment analysis revealed the C365-344 bp fragment had 84% homology with a DNA sequence in a DNA clone, OJ1001_D05, located on chromosome 2 in rice, which had a 19.1 kb distance from two TGMS genes, ptgms2-1 and tms5. In the same time, this fragment also showed as high as 98% homology with a DNA sequence between OJ1001_B06.3 and OJ1001_B06.4 clones located on chromosome 6 in rice, and had about 1 cM distance from a TGMS gene, TMS. The correlation value between the genotypes of C365-1 locus and the fertile phenotypes in 146 individual plants is 0.944, had a highly significant correlation, and each lines of the 15 TGMS lines possessed the same genotype as that of sterile plants DNA pool. The results indicated that PCR marker C365-1 could be used to identify TGMS gene possessed by TGMS plants in rice effectively, which could be an effective marker in TGMS lines marker-assisted selection in rice.