CAJ | 학술논문

黄管秦艽(Gentiana officinalis)是一种重要的藏药高山植物,本研究构建了该物种开花期的cDNA文库.经检测达到中等cDNA文库水平,文库滴度为1.2×107pfu/ml,重组率95.9%,插入片段平均长度大于500bp.对343个随机挑选的重组克隆进行部分测序,获得的ESTs经编辑后共有181条有效序列.经生物信息学方法分析181条表达序列标签(EST)代表144个单克隆序列,其中55个与已鉴定的基因同源,35个序列与未鉴定的EST匹配,54个未找到同源序列;后两者共有89个EST序列未发现功能相似的蛋白.对已鉴定的EST进行功能分析发现,相关基因主要编码以下蛋白:与蛋白表达相关的占35%;光合作用相关的占22%;新陈代谢相关的占18%;抗性相关的占11%;质膜运输和细胞分裂相关的分别占5%;染色体变化和细胞信号转导的分别占2%.根据有效EST序列设计引物,通过RT-PCR进一验证了所得EST的准确性.这些研究结果为将来研究黄管秦艽的功能基因以及该物种与相关物种的群体遗传学、进化生物学等方面提供了基础.
황관진구(Gentiana officinalis)시일충중요적장약고산식물,본연구구건료해물충개화기적cDNA문고.경검측체도중등cDNA문고수평,문고적도위1.2×107pfu/ml,중조솔95.9%,삽입편단평균장도대우500bp.대343개수궤도선적중조극륭진행부분측서,획득적ESTs경편집후공유181조유효서렬.경생물신식학방법분석181조표체서렬표첨(EST)대표144개단극륭서렬,기중55개여이감정적기인동원,35개서렬여미감정적EST필배,54개미조도동원서렬;후량자공유89개EST서렬미발현공능상사적단백.대이감정적EST진행공능분석발현,상관기인주요편마이하단백:여단백표체상관적점35%;광합작용상관적점22%;신진대사상관적점18%;항성상관적점11%;질막운수화세포분렬상관적분별점5%;염색체변화화세포신호전도적분별점2%.근거유효EST서렬설계인물,통과RT-PCR진일험증료소득EST적준학성.저사연구결과위장래연구황관진구적공능기인이급해물충여상관물충적군체유전학、진화생물학등방면제공료기출.
Gentiana officinalis,an alpine plant, one of the widely used Tibetan traditional medicines. In this study, total RNA was extracted from the whole plant of flowering individuals of this species, and a cDNA expression library was constructed using CreatorTM SMARTTM cDNA Library Construction Kit. The results showed that the titer of the cDNA expression library was 1.2×107 pfu/ml and the efficiency of recombination was 95.9%. The average length of insert fragments in the library was longer than 500 bp. A total of 181 valid ESTs were obtained from random sequencing of 343 clones. Further bioinformatic analyses suggested they represented 144 unique clonal sequences in which 55 sequences showed high homology to previously identified genes in Gentianaceae or other plants, 35 sequences matched to other uncharacterized expressed sequence tags (ESTs), and 54 sequences showed no well matches to available sequences in DNA databases. No protein matched to the latter two sorts of ESTs (89). Fifty-five ESTs with matched proteins were involved in a series of diverse functions: protein expression (35%), photosynthesis (22%), metabolism (18%), defense (11%), membrane transport (5%), cell division (5%), chromosome metabolism (2%) and signaling components (2%). At last, RT-PCR primers were designed according to the effective ESTs to amplified the cDNAs of G.officinalis,which further verified the accuracy of the ESTs. This cDNA library provided a critical basis for further analyses of functional genes and gene expression in this alpine species. In addition, these ESTs could be used to design functional nuclear primers for studying population genetics of this species and closely related species.