应用cDNA-AFLP技术分离安吉白茶阶段性返白过程中的差异表达基因,获得一白期表达上调片断TDF（transcript derived fragment, TDF）。BLAST比对结果显示,该片段与其他物种的泛素活化酶基因有很高的相似性。通过SMART-RACE技术分别扩增出其3′和5′末端序列,成功获得该基因全长cDNA序列（GenBank登录号JN180299）。所得序列全长3764bp,其开放阅读框编码1094个氨基酸,蛋白分子量约为121kD。该基因的氨基酸序列与烟草、蓖麻、水稻、小麦、拟南芥中的UBA1基因编码的氨基酸序列分别有82%、81%、79%、79%、77%的同源性。qRT-PCR分析表明,安吉白茶UBA1基因在白化期的表达量是返绿期的2.49倍。泛素活化酶是泛素蛋白酶体介导的蛋白质降解系统中1个关键酶,茶树泛素活化酶基因的克隆为进一步研究安吉白茶阶段性白化的分子机理奠定了基础。
The cDNA-AFLP technology was applied to analyze gene expression during periodic albinism process of Anji Baicha. Some transcript-derived fragments (TDFs) were isolated occurring in both the albinistic and re-greening stage leaves. One of them showed a high similarity to ubiquitin-activating enzyme 1 (UBA1) gene. Based on the fragment, the full length of UBA1 gene with 3764bp (GenBank Accession No. JN180299) cDNA was obtained via rapid amplification of cDNA ends (RACE), named Camellia Sinensis UBA1 gene. It contained an open reading frame (ORF) encoding a polypeptide of 1094 amino acid residues with a predicable molecular mass of 121kD. Analysis of the nucleotide sequence and deduced amino acid sequence showed 82%, 81%, 79%, 79%, 77% homology with UBA1 genes from Nicotiana tabacum, Ricinus communis, Oryza sativa subsp. japonica, Triticum aestivum, Arabidopsis thaliana, respectively. Analysis by qRT-PCR showed that the transcript of UBA1 was significantly up-regulated at the albinistic stage to 2.49-fold higher than that at the re-greening stage. This is a key enzyme in the ubiquitin-proteasome mediated protein degradation system. The clone and analysis of the tea plant UBA1 gene establishes a good foundation for further study on the molecular mechanism of periodic albinism in Anji Baicha.
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