低纬高原紫色辣椒果实花色苷合成相关关键基因的挖掘

Discovery of Fruit Anthocyanin Synthesis-related Key Genes of Purple Chilli in Low Latitude Plateau Area

【目的】挖掘低纬高原生境中紫色辣椒果实花色苷合成相关的关键基因,为紫色辣椒果实花色苷合成分子机理研究和新种质创制奠定基础。【方法】以文山低纬高原主栽且抗性良好的紫色辣椒品种16HN01试材,用色价法和液相色谱串联质谱法分别测定16HN01各发育阶段果实[按长度并结合颜色将果实分为6个发育阶段(1~6),即长度≤1.5 cm、1.5~2.5 cm、2.5~3.5 cm、≥3.5 cm、青熟期与成熟期果实]的总花色苷含量(TAC)和主要花色苷单体含量(AMC),基于转录组测序和qRT-PCR与双因素相关性分析,挖掘果实花色苷合成相关的关键基因。【结果】16HN01果实TAC在发育阶段3最高,为4.626±0.202(A539-0.25A657)/g;主要花色苷单体为飞燕草素-3-O-(2'''-O-对香豆酰)芸香糖苷[Dp-3-O-(2'''-O-C)R]、飞燕草素-3-O-芸香糖苷(Dp-3-O-R)、飞燕草素-3-O-葡萄糖苷(Dp-3-O-G)和飞燕草素-3,5-O-二葡萄糖苷(Dp-3,5-O-DG),其含量分别在阶段3、阶段1、阶段1和阶段3最高;TAC、Dp-3-O-(2'''-O-p-C)R、Dp-3-O-R、Dp-3-O-G和Dp-3,5-O-DG含量与果实中UDPG类黄酮糖基转移酶基因(UFGT)转录水平之间的Pearson相关系数最高,分别为0.848(显著)、0.779、0.308、0.490和0.952(极显著),与WD40转录水平间的相关系数最高,分别为0.977(极显著)、0.702、0.163、0.357和0.674。【结论】低纬高原紫色辣椒果实花色苷合成相关关键结构基因主要包括UFGT、花色素合酶基因(ANS)65、ANS08和查尔酮合酶基因(CHS),转录因子基因WD40和bHLH分别极显著和显著正调控花色苷的合成,MYB和BBX负调控花色苷的合成。

【Objective】The key genes related to the fruit anthocyanin synthesis of purple chilli in the habitat of low latitude plateau were investigated,which laid the foundation for studying the molecular mechanism of the fruit anthocyanin synthesis and the creation of the new germplasm of the purple chilli.【Method】Taking purple chilli 16HN01 with good disease resistance cultivated in Wenshan low latitude plateau area as test material,the total anthocyanin contents(TACs)and main anthocyanin monomer contents(AMCs)of 16HN01 fruits at different developmental stages[according to the length and color,fruits can be divided into six developmental stages(1-6),respectively length≤1.5 cm,1.5-2.5 cm,2.5-3.5 cm,≥3.5 cm,green ripe period and maturity period]were determined by color value method and liquid chromatography-tandem mass spectrometry,respectively,and then the fruit anthocyanin synthesis-related key genes were discovered based on transcriptome sequencing(RNA-seq),qRT-PCR and two factor correlation analysis.【Result】The TAC of 16HN01 fruit was the highest at the Stage 3,which was 4.626±0.202(A539−0.25A657)/g.The main anthocyanins monomers were delphinidin-3-O-(2'''-O-p-coumaroyl)rutinoside[Dp-3-O-(2'''-O-p-C)R],delphinidin-3-O-rutinoside(Dp-3-O-R),delphinidin-3-O-glucoside(Dp-3-O-G)and delphinidin-3,5-O-diglucoside(Dp-3,5-O-DG),and their contents reached the highest at the Stage 3,Stage 1,Stage 1 and Stage 3,respectively.The Pearson correlation coefficients between the contents of the TAC,Dp-3-O-(2'''-O-p-C)R,Dp-3-O-R,Dp-3-O-G and Dp-3,5-O-DG and the transcription level of the UDPG flavonoid glycosyltransferase gene(UFGT)in the fruits were the highest,which were 0.848(significant),0.779,0.308,0.490 and 0.952(extremely significant),respectively,and the correlation coefficients between the contents of the TAC,Dp-3-O-(2'''-O-p-C)R,Dp-3-O-R,Dp-3-O-G and Dp-3,5-O-DG and the WD40 transcription level were the highest,which were 0.977(extremely significant),0.702,0.163,0.357 and 0.674,respectively.【Conclusion】The key structural genes related to the fruit anthocyanin synthesis of purple chilli in the low latitude plateau area mainly include UFGT,anthocyanin synthase gene(ANS)65,ANS08 and chalcone synthase gene(CHS).Transcription factor genes WD40 and bHLH are very significantly and significantly positively regulated the anthocyanin synthesis,whereas MYB and BBX are negatively regulated the anthocyanin synthesis.