棉花育性相关基因GhPG2的克隆与表达分析

doi:10.3724/SP.J.1006.2009.02008

摘要/Abstract

摘要：

多聚半乳糖醛酸酶家族是一个细胞壁水解酶，在许多高等植物的花粉中都具有高水平外切多聚半乳糖醛酸酶(PG)活性，可能在花粉成熟和花粉管伸长中发挥重要作用。利用已报道与恢复基因连锁的分子标记STS679筛选陆地棉Y18恢复系核基因组BAC文库，获得一个阳性克隆Z55E7，对该克隆测序获得6个基因，其中一个为编码多聚半乳糖醛酸酶(polygalacturonase)基因，暂被命名为GhPG2。在棉花细胞质雄性不育系、保持系和恢复系中对该基因进行序列和表达分析，以便进一步了解该基因与棉花细胞质雄性不育的关系。从BAC克隆中分离到GhPG2基因，对其进行序列分析，以半定量RT-PCR和实时定量PCR分析该GhPG2基因在棉花恢复系、保持系和不育系各个组织中的表达模式。分离到GhPG2基因的基因组序列为1 518 bp，其开放阅读框序列为1 239 bp，共编码412个氨基酸，包含PG基因家族的4个保守结构域。系统进化树分析表明，GhPG2基因与其他物种花粉表达的PG基因同属C分支。表达分析表明, 该基因在恢复系和保持系的花药、花瓣和VI级花蕾中高表达，而不育系的相应组织该基因表达量明显降低。GhPG2基因归类于花粉表达的一类PG基因。实验结果暗示GhPG2基因可能与棉花正常的花器官发育相关。

关键词: 棉花, 多聚半乳糖醛酸酶, GhPG2, 花粉发育, 表达谱分析

Abstract:

Polygalacturonase (PGs) gene family is a kind of proteolytic enzyme in cell wall. A number of higher plants have a higher level of the excision activity at pollen. Polygalacturonase plays an important role on the processes of pollen mature and pollen tube growth. Through BAC library screening, a BAC clone Z55E7 was obtained and congregated with molecular marker STS679 by using PCR strategy. Sequencing analysis results showed that there were six genes in the clone. One of them was a Polygalacturonase gene in cotton, named GhPG2. The expression profiles analysis of this gene would help to know the relationshipof GhPG2 gene with the molecular mechanism of cotton cytoplasm male sterile. GhPG2 gene was isolated from BAC clone. The amino acids and nucleotide sequence of the gene were analyzed by bioinformation software. GhPG2 gene expression levels of different tissues in CMS line, maintainer and restorer line were analyzed by RT-PCR and real-time quantitative PCR method. The gene with the sequence of 1518 bp and an open reading frame of 1239bp coded 412 amino acids and included four conserved domains. Multiple alignment and phylogenetic analysis based on the amino acid sequences showed that GhPG2 was categorized as a gene expressed in pollen. GhPG2 gene expression level in anther, petal and the sixth grade flower bud was higher in the maintainer and restorer lines and lower obviously in the CMS line. And its relative transcript levels were 1.73, 1.45; 3.7, 3.4; 1.63, and 1.39 folds compared with petal, anther and VI flower bud of sterile line, respectively.The gene expression level in anther, petal and the sixth grade flower bud was no significance between the maintainer and restorer line with relative transcript levels were 4.33, 4.08; 13.29, 12.12; 3.25 and 3.10 folds, respectively. It is speculated that GhPG2 gene is of significance in floral organ development in cotton.

Key words: Cotton, GhPG2, Flower Development, Polygalacturonase, Expression profiling

侯思宇，张锐，郭三堆. 棉花育性相关基因GhPG2的克隆与表达分析[J]. 作物学报, 2009, 35(11): 2008-2014.

HOU Si-Yu,ZHANG Rui,GUO San-Dui. Cloning and Expression Profile Analysis of Ghpg2 Gene Associated with Fertility in Cotton（Gossypium hirsutum L.）[J]. Acta Agron Sin, 2009, 35(11): 2008-2014.

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