不同颜色刺葡萄花色苷合成相关基因的表达分析

Analysis on anthocyanin profiles and expressions of related biosynthesis structural genes in berry skin of Vitis davidii

【目的】探索不同果皮颜色中国野生刺葡萄(Vitis davidii)花色苷合成相关基因的表达差异,为中国野生刺葡萄资源的利用奠定基础。【方法】以白色及黑色果实的中国野生刺葡萄、欧亚种葡萄‘白比诺’及‘黑比诺’为材料,利用HPLC-ESI-MS/MS分析不同发育期果皮中花色苷的组成及其含量,通过序列检测分析几种刺葡萄Vvmy-bA1基因序列的不同,用实时荧光定量PCR检测花色苷合成相关结构基因表达的差异。【结果】锦葵色素葡萄糖苷(Malvidin 3-O-glucoside)仅在黑色果实转色后被检测到,其在中国野生刺葡萄黑色果实和‘黑比诺’中的含量分别为0.12～7.14mg/kg与4.90～180.79mg/kg,其他花色苷如矢车菊素3,5-O-双葡萄糖苷(Cyanidin 3-O-glucoside-5-O-glucosid)、飞燕草素3-O-葡萄糖苷(Delphinidin 3-O-glucoside)、花葵素3-O-葡萄糖苷(Pelargonidin 3-O-glucoside)、矮牵牛色素3-O-葡萄糖苷(Petunidin 3-O-glucosid)等在黑、白色葡萄果实不同发育期均能检测到,其含量为0.01～49.28mg/kg。VvmybA1a等位基因在中国野生刺葡萄黑、白色果实中都存在,且序列与欧亚种一致;VvmybA1c等位基因只在中国野生刺葡萄黑色果实中可检测到,与欧亚种相比其序列有33bp的插入片段和47bp的缺失片段。My-bA1基因只在黑色果实成熟期表达;合成花色苷的关键结构基因UFGT在供试葡萄各个时期均有表达,但其表达量在黑色果实成熟期显著高于白色果实;F3′5′H、GST及OMT等结构基因也在黑色果实成熟期表达上调,而其他结构基因的表达则在中国野生刺葡萄和欧亚种之间表现出了较大差异。【结论】中国野生刺葡萄花色苷合成相关基因具有特殊结构,与欧亚种葡萄相比有明显差异。

【Objective】This study investigated the differences in relative expression of anthocyanins biosynthesis structural genes between white skin berry and corresponding colored type of Chinese wild grape V.davidii to provide basis for its utilization in future.【Method】The compositions and contents of anthocyanins from four cultivars/lines including white skin berry and colored berry of V.davidii,‘Pinot Blanc’and‘Pinot Noir’of V.vinifera were analyzed by HPLC-ESI-MS/MS.The MybA1 transcription factors regulating anthocyanin biosynthesis were detected,and the gene sequences of MybA1 were cloned from different samples and compared to explore the relationships between Vitis species.The expressions of anthocyanins related biosynthesis structural genes were analyzed by real-time PCR.【Result】Only malvidin 3-Oglucoside was detected in color skin grapes during ripening stage,and the contents of colored V.davidii and‘Pinot Noir’were 0.12-7.14 mg/kg and 4.90-180.79 mg/kg,respectively.The contents of other anthocyanins(0.01-49.28 mg/kg),such as cyanidin 3-O-glucoside-5-O-glucosid,delphinidin 3-O-glucoside,pelargonidin 3-O-glucoside and petunidin 3-O-glucosid,were lower by several-fold to hundreds-fold compared to the colored grape varieties.Compared with V.vinifera,there were a 33 bp short insertion and a 47 bp gene fragment missing in the VvmybA1 c allele of V.davidii.MybA1 regulated anthocyanin accumulation in grape berry skin via UFGT gene expression.Colored berry skin had high levels of MybA1 and UFGT genes expression at harvest stages compared with white berry skin.In addition,the levels of F3′5′H,GSTand OMTgenes expression were similar to UFGT.Other structure genes expressions were significantly different between V.davidii and V.vinifera.【Conclusion】There were significant differences identified in the sequences of MybA1 between V.viniferaand V.davidii.