甘蓝型油菜光叶性状的遗传分析及基因定位

Genetic Analysis and Gene Mapping of Glossy Leaf in Brassica napus

【目的】表皮蜡质是覆盖植物叶片和茎秆等部位的疏水层,在植物抵御逆境方面发挥着重要作用,同时会影响光合作用和生长发育;甘蓝型油菜作为世界上主要的油料作物之一,研究表皮蜡质突变体的遗传机制,为实现油菜高产稳产提供参考。【方法】对油菜光叶突变体M8和普通蜡质叶中双11(ZS11)和C20的叶片表征进行记载,使用便携式植物光合作用测量系统测定叶片光合速率;利用M8和ZS11、C20分别杂交获得2个F1、自交构建2个F2分离群体,用于分析油菜光叶性状的遗传规律;选取M8和ZS11杂交的F2群体中光叶和蜡质叶表型单株分别进行混池,通过集团分离分析法结合靶向测序技术进行基因克隆,结合比较基因组和转录组数据进行候选基因预测,并通过RT-PCR验证候选基因。【结果】甘蓝型油菜光叶表型叶片气孔导度更大、光合效率更高;遗传分析表明M8光叶性状受1对基因控制,光叶相对蜡质叶为隐性。图位克隆将光叶控制基因定位至A08染色体0.134—0.699 Mb物理区间内。进一步分析发现,相比于ZS11,光叶突变体M8中A08染色体0.22—0.58 Mb区间存在大片段缺失,ZS11在该区段中的Bna A08G0006900ZS(Bna A08.SAGL1)可能为光叶的候选基因,该基因编码一个Kelch-F-box蛋白,在ZS11叶片中表达量高而M8中未检测到,该基因缺失可能导致了M8光叶表型。【结论】甘蓝型油菜光叶新突变体M8相比野生型蜡质叶片光合速率更高,该光叶表型受1对隐性基因控制;图位克隆鉴定到光叶性状调控基因为Bna A08.SAGL1,基因缺失产生了光叶表型。

【Objective】Epicuticular waxes is a hydrophobic layer covering plant leaves and stems,playing an important role in stress resistance and affecting photosynthesis,growth and development.Rapeseed(Brassica napus L.)is one of the main oil crops in the world,but the mechanism of epicuticular waxes formation on its leaves is still unclear.This study aims to reveal the genetic mechanism of epicuticular wax on leaves through a mutant,which will help to achieve high and stable yield of rapeseed.【Method】In this study,leaf of glossy leaf mutant M8 and common waxy leaf inbred lines Zhongshuang 11(ZS11)and the C20 were characterized,and its photosynthetic rate were measured by portable plant photosynthesis measurement system.Two F1 hybrids and F2 segregating populations were constructed by crossing the M8 with ZS11 and C20,respectively,and were used to analyze the heredity of leaf characters in rapeseed.The F2 population hybridized by M8 and ZS11 was constructed to analyze the genetic regulation of the glossy leaf trait in rapeseed by bulked segregation analysis(BSA)combined with Target-sequencing(Target-seq).Combined with comparative genome and transcriptome database, candidate genes were predicted and then verified by RT-PCR.【Result】The leaf stomatal conductance and photosynthetic efficiency were higher in the glossy leaf mutant M8, compared with ZS11 in rapeseed. The genetic analysis results showed that the glossy leaf was controlled by one pair of recessive genes and glossy leaf was recessive compared with waxy leaf. It was finally fine mapped in the physical region of 0.134-0.699 Mb on chromosome A08 by map based cloning method. Further analysis revealed that there was a large segmental deletion in the 0.22-0.58 Mb region of chromosome A08 in the M8 glossy leaf mutant compared to ZS11, and BnaA08G0006900ZS (BnaA08.SAGL1) in this region of the genome was identified as the candidate gene for the glossy leaf trait. This gene encodes a Kelch-F-box protein, and its deletion may lead to the observed glossy leaf trait in the M8 mutant, as it was highly expressed in ZS11 but none in M8. 【Conclusion】Compared with wild-type waxy leaves, the photosynthetic rate of the new glossy leaf mutant M8 was higher, and the glossy leaf phenotype was controlled by a recessive gene. In the picture, the glossy leaf phenotype regulatory gene was identified as BnaA08.SAGL1, and deletion of the gene resulted in glossy leaf in rapeseed.