CAJ | 학술논문

该研究对黄灯笼椒 pal基因进行了PCR扩增、测序及生物信息学分析，在此基础上构建原核表达载体（pET-32a-pal）进行原核表达,分析外施茉莉酸条件下基因表达模式和辣椒素积累情况。生物信息学分析表明黄灯笼椒 pal基因编码的蛋白质由683个氨基酸残基组成，蛋白质的分子量为74.28 kD，等电点为6.97。将原核表达载体在大肠杆菌 E. coli BL21中进行诱导表达，SDS-PAGE分析表明，诱导后表达目的蛋白质相对标准分子质量约为74 kD，与 DNAMAN软件预测的（74.2 kD）基本一致。蛋白质序列比对和进化树分析表明，黄灯笼椒 Capal蛋白与朝天椒 Capal蛋白一致性较高，进化距离最近。Real-time PCR分析表明，外源茉莉酸处理对 pal基因表达有上调作用；高效液相色谱（HPLC）分析茉莉酸处理后辣椒素含量发现，外源茉莉酸可以促进辣椒素的合成。该研究将为 pal基因转录调控和功能研究提供参考。
해연구대황등롱초 pal기인진행료PCR확증、측서급생물신식학분석，재차기출상구건원핵표체재체（pET-32a-pal）진행원핵표체,분석외시말리산조건하기인표체모식화랄초소적루정황。생물신식학분석표명황등롱초 pal기인편마적단백질유683개안기산잔기조성，단백질적분자량위74.28 kD，등전점위6.97。장원핵표체재체재대장간균 E. coli BL21중진행유도표체，SDS-PAGE분석표명，유도후표체목적단백질상대표준분자질량약위74 kD，여 DNAMAN연건예측적（74.2 kD）기본일치。단백질서렬비대화진화수분석표명，황등롱초 Capal단백여조천초 Capal단백일치성교고，진화거리최근。Real-time PCR분석표명，외원말리산처리대 pal기인표체유상조작용；고효액상색보（HPLC）분석말리산처리후랄초소함량발현，외원말리산가이촉진랄초소적합성。해연구장위 pal기인전록조공화공능연구제공삼고。
The cDNA sequence of Capal gene was cloned from Capsicum chinense Jacq by RT-PCR and sequenced. Bioinformatics analysis showed that Capal en-codes a putative polypeptide of 683 amino acids with a calculated molecular mass of 74.2 kD and a theoretical pI of 6.9. Multiple sequence alignments and phyloge-netic tree analyses showed that Capal protein of C. chinense is similar to that of Capsicum annuum var. conoides, with an overal sequence similarity of 96%. The prokaryotic expression vector pET-32a-pal was constructed and induced to express in E. coli BL21. The SDS-PAGE analysis showed that the relative molecular mass of the induced new protein is about 74 kD, which was basical y identical with that predicted by DNAMAN software (74.3 kD). Real-time PCR analysis showed that ex-ogenous jasmonic acid (JA) promoted pal expression. The accumulation of capsaicin in pepper was analyzed by high performance liquid chromatography (HPLC), and the results indicated that exogenous jasmonic acid (JA) can promote the synthesis of capsaicin. This study wil provide valuable experimental basis for the research of transcription regulation and explaining the gene function of pal.