目的 克隆寧夏枸杞類黃酮/異黃酮生物合成途徑中的關(guān)鍵酶查耳酮合成酶（CHS，EC2.3.1.74）。方法 以寧夏枸杞為材料，利用同源克隆和RT-PCR技術(shù)得到1條CHS基因，并對其進行生物信息學分析。結(jié)果 該家族的cDNA 片段長為1 148 bp，編碼382個氨基酸，蛋白相對分子質(zhì)量為4.168×10⁴，等電點為6.22。氨基酸序列和結(jié)構(gòu)分析顯示CHS基因家族有一個活性位點，即查耳酮和二苯乙烯合成酶活性位點。寧夏枸杞CHS（LyCHS）蛋白很可能定位在細胞質(zhì)。對二級結(jié)構(gòu)和三級結(jié)構(gòu)進行預測，發(fā)現(xiàn)LyCHS蛋白中無規(guī)則卷曲126個，α螺旋和β折疊分別為166和25個，延伸鏈65個。系統(tǒng)進化分析表明，LyCHS基因與馬鈴薯野生種、馬鈴薯、番茄具有很高的同源性。該基因已在GenBank上注冊，基因序列登錄號為JQ964237。結(jié)論 首次克隆了LyCHS基因片段，為研究其表達特性以及功能提供了基礎(chǔ)。

Objective To obtain cDNA of chalcone synthase (CHS, EC 2.3.1.74) involved in the flavonoid/isoflavonoid biosynthesis pathway. Methods The partial sequence of CHS in Lycium barbarum (LyCHS) was successfully cloned by RT-PCR and using a sequence homology strategy, and the bioinformation analysis was carried out. Results The cDNA fragment of CHS gene was 1 148 bp in length, which encoded a protein of 382 amino acids with the predicted relative molecular weight of 4.168 × 10⁴. The estimated isoelectric point (pI) of the putative protein is 6.22. According to the amino acid sequence and structural analysis, it showed that this protein contained one conserved active site, namely chalcone and stilbene synthases active site. Subcellular localizations of LyCHS proteins were likely in the cytoplasm. The secondary and tertiary structures of LyCHS were abundant in α-helixs (166) and random coils (126), while were less in β-turns (25) and extended strains (65). Phylogenetic analysis showed that the genes of LyCHS were closely related to CHS in Solanum pinnatisectum, S. tuberosum, and Lycopersicon esculentum. The sequences had been registered in GenBank with the accession numbers JQ964237. Conclusion The present results provide the foundation for the next study of CHS gene about its expression and function in L. barbarum.

北方民族大學校級科研項目（2010Y046）