环境因子对“紫娟”茶树叶片呈色与花青素积累效应影响的研究

“Zijuan”, a specific tea variety from breeding of Tea Research Institute，Yunnan Academy of Agricultural Sciences, has rather high drinking and medical value. With the expanding of planting area, it is found that "Zijuan” cultivating was area restriction, Therefore, there is scientific significance and practical value to study effect of environmental factors on leaf coloring and anthocyanin accumulation of tea cultivars "Zijuan". In order to ensure the inherently quality of "Zijuan"tea and benefit of consumer and producer. In this project, the transcriptome of leaf of “Zijuan” from two different cultivation area is sequenced through RNA-seq technology, Firstly, it would screen key genes related to leaf color after comparing coloring metabolic pathways of leaf color from two transcriptome. Secondly, the full-length cDNA of those genes would be amplified, and the spatiotemporal expression of genes would be studied by quantitative RT-PCR. Thirdly, the effect of expression genes and anthocyanin accumulation by environmental factors (light, temperature, fertilized, hormone) would be also determined. Through these works, it would not only clarify effect of environmental factors on leaf coloring and anthocyanin accumulation of tea cultivars "Zijuan", and provide a theoretical basis for choosing suitable cultivation area and management measures of high anthocyanin in “Zijuan” tea.

“紫娟”是云南省农业科学院茶叶研究所选育的特异茶树品种，具有较高的饮用和药用价值。随着种植区域和面积的扩大，发现“紫娟”茶树的种植受区域限制。因此，为了保证“紫娟”茶应有的质量以及消费者和生产者的利益，开展环境因子对“紫娟”茶树叶片呈色以及花青素积累的调控具有重要的科学意义和实用价值。本项目采用RNA-seq技术对来至两个不同栽培区的“紫娟”幼嫩芽叶进行转录组测序，一是比较分析两个样本转录组中有关色泽代谢相关途径，筛选出重要的呈色关键基因；二是对重要呈色基因利用RACE技术进行全长cDNA克隆以及利用荧光定量PCR研究其在“紫娟”茶树中的时空表达特征；三是研究环境因子（光照、温度、施肥、激素）对呈色相关基因的表达以及花青素积累调控的影响。通过上述研究，明确外界因子对“紫娟”茶树叶片呈色以及花青素积累的影响，为“紫娟”茶树栽培适宜区域的推广和高花青素的“紫娟”茶园管理措施提供理论依据。

“紫娟”是珍稀茶树种质的典型代表，其叶片色泽呈现紫色，富含花青素、儿茶素以及黄酮等活性成分。为探讨外界环境因子对紫娟茶树叶片呈色与花青素积累的影响，本课题选用叶色为绿色且花青素含量低的茶树为对照，通过RNA-seq、数字基因表达谱，并结合生物信息学等方法与技术，获得166 118条Unigene以及鉴定出46种已知的miRNAs和67种新的miRNAs，筛选出与紫娟叶片呈色的关键基因9个（PAL、C4H、4CL、CHS、DFR、ANS、UFGT以及FLS、ANR）和参与调控花青素合成miRNA 4个（miR828a、miR845c、novel_14和novel_87）；通过对紫娟叶片的全长转录组测序，获得具有完整编码的CDS 199 245个，包含花青素合成的关键基因；通过对不同pH和缺素（Fe、K、Mg、N、P、Zn、Mn）条件下花青素组分分析以及花青素合成关键基因表达分析，初步明确“紫娟”的最适生长pH 为4.5-6.0，缺素（Fe、K、Mg、N、P）条件下，“紫娟”新梢中花青素含量增加，ANS、UFGT为其响应的关键基因；缺（Zn、Mn）处理花青素含量降低，同时发现蓝光能诱导“紫娟”花青素合成相关基因的表达，这些说明土壤pH、土肥条件和光照对“紫娟”花青素的积累具有重要的调控作用。上述研究结果为后续基因功能验证以及阐明“紫娟”茶树叶片的呈色机制奠定研究基础。通过项目实施发表科技论文8篇，2名科技人员晋升研究员。