关键词: 茶树, 咖啡碱, 次黄嘌呤核苷酸脱氢酶, TIDH, 基因克隆, 关联分析

Abstract: Caffeine is one of the most important purine alkaloids，as it plays an important role in
flavor of tea. It’s of great importance to study the caffeine biosynthesis pathway of tea plant. The primers
were designed based on TIDH（which codes for inosine-5'-monophosphate dehydrogenase，a key gene in
the caffeine biosynthesis pathway of tea plant）cDNA from the NCBI. One internal fragment of TIDH
genomic DNA was cloned（4 172 bp，TIDH3）. A set of 95 accessions which are typical in caffeine content
was selected from the core collection of Chinese tea germplasm previously established. The caffeine
content of two seasons of spring and autumn in two years was tested by HPLC. The result showed the
caffeine content of most germplasms was 2.50%–4.50%，the average content was 3.50%. The coefficient
of variation is 15.52%–19.25%，which means that the caffeine content of tea is stable between different
years or seasons. A 759 bp genomic DNA fragment of TIDH3 was cloned，sequenced and analyzed. The
average nucleotide diversity for the sequenced region was calculated to be πT = 0.06 and θW = 0.012. The
diversity at synonymous sites，π_syn = 0.009，was higher than diversity at non-synonymous sites，πnonsyn =
0.006. The ratio of Ka/Ks was 0.600（lower than 1.0），indicating the action of purifying selection. The
linkage disequilibrium（LD）of SNPs in TIDH3 was detected and the result showed that LD declined
rapidly within the gene region（the value of r2 reduced to 0.2 in 300 bp）. By means of association analysis，
two SNPs were found to be significantly correlated with caffeine content，their genetic effect values were
10.43% and 5.68%，respectively.

Key words: tea plant, caffeine, TIDH, gene cloning, association analysis