摘要
棉纤维正常发育需要大量的蔗糖供应。转化酶是生物体内蔗糖代谢途径中的关键酶之一,对转化酶基因的结构和功能研究将有助于揭示复杂的棉纤维发育分子机制,并为纤维品质改良提供优良的基因资源。本研究以棉纤维突变体im和遗传标准系TM-1纤维发育中显著差异表达的EST序列(GenBank登录号为EY196825)为探针,通过电子克隆方法,结合cDNA及基因组全长基因PCR扩增、测序验证,克隆到一个棉花液泡转化酶基因GhVacInv2a(GenBank登录号为KF305322)。该基因ORF全长1857 bp,编码618个氨基酸,与已登录的陆地棉GhVacInv2基因(GenBank登录号为FJ864677)序列一致性为99%(E=0)。基因组序列分析表明,GhVacInv2a在二倍体棉种非洲棉和雷蒙德氏棉中含1个拷贝,在四倍体陆地棉和海岛棉中存在2个拷贝,其中GhVacInv2a和GhVacInv2分属于A、D亚组同源基因。该同源基因含7个外显子,6个内含子。Q-PCR分析表明,该基因在花药中表达量最高,在快速伸长的纤维组织中优势表达。在13～19 DPA的纤维中,其在TM-1中的表达水平极显著地高于im突变体。开发SNP标记,将GhVacInv2a基因定位在异源四倍体棉花第3染色体上。标记与性状的关联分析显示,GhVacInv2a与纤维强度存在显著相关(P=0.0087),表明GhVacInv2a在棉花纤维品质形成中可能发挥重要功能。
Cotton fiber development depends on a large supply of sugar. Invertase plays a central role in sucrose metabolism. Structural and functional analysis of gene encoding invertase will conductively reveal the complex molecular mechanism of fiber development, and also provide elite gene resources for improvement of cotton fiber quality. In the study, a novel gene encoding vacuolar invertase was obtained by sillico cloning, based on a differentially expressed EST （GenBank accession number EY196825） between G. hirsutum acc. TM-1 and im mutant, combined with reconfirmation of ORF information in transcriptional and genomic level. BlastN showed that the gene shared 99% identity of GhVacInv2 reported previously （GenBank accession number FJ864677）, and designated as GhVacInv2a （GenBank accession number KF305322）. GhVacInv2a contained an open reading frame of 1857 bp that encoded a polypeptide of 618 amino acids, with seven exons and six introns at genome sequence. GhVacInv2a comprised one copy in diploid cotton species G. herbaceum and G. raimondii, and two copies in tetraploid cotton species G. hirsutum acc. TM-1 and G. barbadense cv. Hai 7124, with GhVacInv2a in A-subgenome and GhVacInv2 in D-subgenome, respectively. Q-PCR expression analysis showed that the set of homoelogous genes performed the highest expressive abundance in anther in comparison with other tissues. In developing fibers, it was predominantly expressed from fiber rapid elongation stage, and there existed significant difference between TM-1 and im mutant during 13 to 19 days post anthesis （DPA） in fiber tissues. Further, GhVacInv2a was located on chromosome 3 by developing subgenome-specific SNP marker, and the association analysis showed that there was a significant correction （P=0.0087） between GhVacInv2a and fiber strength, suggesting that GhVacInv2a probably plays a key role in fiber qualities formation.
出处
《作物学报》
CAS
CSCD
北大核心
2014年第3期390-396,共7页
Acta Agronomica Sinica
基金
国家自然科学基金项目(30871558)
江苏省农业科技自主创新资金项目[cx(13)3059]资助
关键词
棉花
纤维发育
液泡转化酶2
克隆
功能分析
Cotton Fiber development Vacuolar invertase 2 Cloning Functional analysis
