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植物LIM蛋白参与细胞骨架的调节及调控次生细胞壁的发育。本研究在雷蒙德氏棉(D5)、亚洲棉(A2)基因组和陆地棉的A、D亚基因组中分别鉴定出14个LIM基因。陆地棉GhLIM基因在2个亚组间的染色体上的分布及基因的复制模式是一致的。推测其复制发生在其祖先同可可(6个LIM)分开之后,A2、D5基因组分化之前;陆地棉LIM除多倍体化使其数目加倍外,没有基因复制与丢失事件发生。蛋白系统进化分析表明,所有被子植物LIM蛋白可以划分为4个亚组(部分具有小亚组):αLIM1(FLIM1、WLIM1、PLIM1)、βLIM1、γLIM2(WLIM2)、δLIM2(PLIM2-I、PLIM2-II)。陆地棉GhLI M的同源复制基因分别在WLIM2和PLIM2-I、PLIM2-II中聚为独立的分支。通过分析深度测序表达谱,发现陆地棉GhLIM有3种表达类型:组成型表达的GhLIM1、2(FLIM),GhLIM3(WLIM1),GhLIM13(βLIM1)和GhLIM4、8、14(WLIM2);雄蕊中优势表达的GhLIM7、9(PLIM2-I),GhLIM5、6、10、11(PLIM2-II);在雄蕊和纤维中优势表达的GhLIM12(PLIM2-I)。实时荧光定量PCR分析表明,GhLIM基因各成员在不同的纤维发育阶段具有不同的表达特征,GhLIM1、13、14在纤维的快速伸长期(5~15 DPA)上调表达,在15 DPA表达量最高;GhLIM2则在纤维次生壁发育期(18~28 DPA)显著上调表达,在28 DPA表达量最高。而GhLIM12在10~23 DPA持续高表达,在28 DPA则没有表达。上述结果说明GhLIM家族基因在纤维发育的不同阶段发挥着不同的功能。
Plant LIM protein is involved in the regulation of cytoskeleton and regulation of secondary cell wall development. In this study, 14 LIM genes were identified in the A, D subgenomic genomes of two cotton cultivars, namely, cotton (D5), Asia (A2) and Upland cotton. The GhLIM gene in upland cotton distributes in the chromosome between two subgroups and the pattern of gene duplication is consistent. It is hypothesized that their replication occurred prior to A2, D5 genomic differentiation after their ancestors were separated from cocoa (6 LIMs); there was no occurrence of gene duplication and loss events until Upland cotton LIM doubly multiplied its number. Protein phylogenetic analysis showed that all angiosperm LIM proteins could be divided into 4 subgroups (some with small subgroups): αLIM1 (FLIM1, WLIM1, PLIM1), βLIM1, γLIM2 (WLIM2), δLIM2 (PLIM2- II). Homologously replicating genes from upland cotton GhLI M cluster into separate branches in WLIM2 and PLIM2-I, PLIM2-II, respectively. By analyzing the deep sequencing expression profiles, we found three expression types of GhLIM: GhLIM1,2 (FLIM), GhLIM3 (WLIM1), GhLIM13 (βLIM1) and GhLIM4,8,14 (WLIM2) GhLIM7,9 (PLIM2-I), GhLIM5,6,10,11 (PLIM2-II); GhLIM12 (PLIM2-I) predominantly expressed in stamens and fibers. Real-time PCR analysis showed that the members of GhLIM gene had different expression characteristics at different stages of fiber development. GhLIM1, 13 and 14 were up-regulated in rapid elongation (5 ~ 15 DPA) and highest in 15 DPA. GhLIM2 was significantly up-regulated during secondary fiber wall development (18-28 DPA) and highest at 28 DPA. However, GhLIM12 continued to be highly expressed at 10 ~ 23 DPA and not expressed at 28 DPA. The above results indicate that GhLIM family genes play different roles in different stages of fiber development.