Molecular cloning and functional analysis of the root-knot nematode resistant gene Me3 in pepper (Capsicum annuum L.)
Molecular cloning and functional analysis of the root-knot nematode resistant gene Me3 in pepper (Capsicum annuum L.)
复制标题
辣椒根结线虫抗性基因Me3的分子克隆及功能分析
DOI：
10.1016/j.hpj.2022.12.003
复制
发表时间：
2022
影响因子：
5.7
通讯作者：
Mao ZC
中科院分区：
文献类型：
--
作者：
Liu Y;Cao HY;Ling J;Yang YH;Li Y;Xie BY;Zhao JL;Mao ZC
辣椒根结线虫是世界范围内辣椒的主要病害之一。在辣椒中，Me 3基因提供了对RKN的热稳定和广谱抗性。本研究利用简单重复序列（SSR）标记和插入/缺失（InDel）标记对Me 3基因进行精细定位。对2272个F_2代个体的分析表明，Me_3位于SSR标记784和SSR标记339之间的45-kb DNA区域内，该区域内有3个候选基因。其中，辣椒品系“HDA 149”的Capana 09 g 000163基因是一个新的核苷酸结合位点和亮氨酸丰富重复序列（NBS-LRR）家族基因，其DNA序列长6348 bp，开放阅读框为2802 bp，编码933个氨基酸，包括NB-ARC和LRR结构域。烟草瞬时表达试验表明Capana 09 g 000163的表达在烟草本氏叶中引发过敏反应（HR）。亚细胞定位结果显示Capana 09 g 000163蛋白定位于细胞核。Capana 09 g 000163在拟南芥中的异位表达与野生型（WT）拟南芥相比显著提高了对南方根结线虫的抗性。此外，M. incognita几乎不能在表达Capana 09 g 000163的转基因拟南芥中发育。因此，本研究利用图位克隆和转基因技术克隆了Me 3基因，并验证了Me 3基因能够诱导辣椒对南方根结线虫的抗性，这对辣椒抗根结线虫育种具有重要意义。
Root-knot nematodes (RKNs) cause severe diseases in peppers annually around the world. In pepper, theMe3gene provides a heat-stable and broad-spectrum resistance to RKNs. In this study, several simple sequence repeat (SSR) markers and insertion/deletion (InDel) markers were developed to fine map theMe3gene. Analysis of 2272 individuals (F2progenies) revealed thatMe3was located in a 45-kb DNA region between markers SSR784 and SSR339, in which there were three candidate genes. Among them, as a novel nucleotide binding site and leucine rich repeat (NBS-LRR) family gene, the DNA sequence ofCapana09g000163of pepper line ‘HDA149’ was 6348 bp in length, with a 2802-bp open reading frame encoding 933 amino acids, including NB-ARC and LRR domains. Tobacco transient expression assays demonstrated that expression ofCapana09g000163triggered a hypersensitive response (HR) inNicotiana benthamianaleaves. Subcellular localization results showed that the Capana09g000163 protein was localized in the cell nucleus. Ectopic expression ofCapana09g000163inArabidopsissignificantly increased resistance againstMeloidogyne incognitacompared with the wild-type (WT)Arabidopsis. Furthermore,M.incognitawas almost unable to develop in transgenicArabidopsisexpressingCapana09g000163. Taken together, we cloned theMe3gene and verified that it induced resistance againstM.incognitawith the methods of map-based cloning and transgenic technology, which may be of great significance to pepper breeding for resistance against RKNs.