CAJ | 학술논문

为板栗 FLC 同源基因控制开花的相关功能及春化作用的相关分子机理提供研究基础，以罗田板栗为研究材料，分离并鉴定板栗花芽调控相关基因 FLC，依据 NCBI 数据库提供的 EST 序列片段，利用RACE 技术，从板栗叶片总 RNA 中分离了与开花相关的基因-CmFLC cDNA 的序列，通过在线分析预测其序列同源性及3D 结构，以 pBluescript SK plus 作为过渡载体构建其干扰表达载体。结果表明，该序列全长为968 bp，其中有1个编码含227个氨基酸蛋白序列的开放阅读框。CmFLC 蛋白的理论等电点为5．80，分子质量为25．62 kDa，N 端含有 M 盒保守序列，该蛋白的二级结构大部分由无规则卷曲和α-螺旋构成。Cm-FLC 存在 M 盒、K 盒两个特征性序列区域。CmFLC 蛋白与毛白杨 FLC 蛋白的三维结构存在高度相似性。板栗的 FLC 蛋白属于植物进化分支，且与山核桃的 FLC 蛋白同属一支。设计特异引物扩增得到 FLC 基因的正、反义基因干扰片段，酶切结果显示插入片段大小为320 bp，板栗 FLC 的干扰载体 pc1301-ubi-CMFLC-RNAi 构建成功。结论：板栗中存在 FLC 同源基因，并可能参与板栗开花相关功能及春化调控。
위판률 FLC 동원기인공제개화적상관공능급춘화작용적상관분자궤리제공연구기출，이라전판률위연구재료，분리병감정판률화아조공상관기인 FLC，의거 NCBI 수거고제공적 EST 서렬편단，이용RACE 기술，종판률협편총 RNA 중분리료여개화상관적기인-CmFLC cDNA 적서렬，통과재선분석예측기서렬동원성급3D 결구，이 pBluescript SK plus 작위과도재체구건기간우표체재체。결과표명，해서렬전장위968 bp，기중유1개편마함227개안기산단백서렬적개방열독광。CmFLC 단백적이론등전점위5．80，분자질량위25．62 kDa，N 단함유 M 합보수서렬，해단백적이급결구대부분유무규칙권곡화α-라선구성。Cm-FLC 존재 M 합、K 합량개특정성서렬구역。CmFLC 단백여모백양 FLC 단백적삼유결구존재고도상사성。판률적 FLC 단백속우식물진화분지，차여산핵도적 FLC 단백동속일지。설계특이인물확증득도 FLC 기인적정、반의기인간우편단，매절결과현시삽입편단대소위320 bp，판률 FLC 적간우재체 pc1301-ubi-CMFLC-RNAi 구건성공。결론：판률중존재 FLC 동원기인，병가능삼여판률개화상관공능급춘화조공。
To provide the research foundation for molecular mechanism of homologous gene FLC controlling flowering-related functions and vernalization of C.mollissima,based on the sequence of EST from NCBI,the authours isolated a flowering locus C gene from Luotia C.mollissims according to RACE technology.The sequence homology and three dimensional structure was analyzed on line, and the interference expression vector was constructed with transition carrier pBluescript SK plus. Results:CmFLC cDNA sequence was 968 bp containing an open reading frame(ORF),which encoded 227 amino acids with a predicted molecular mass of 25.62 kDa and the theoretical isoelectric point(PI)of 5.80. CmFLC was an intro-free gene,and its deduced polypeptide contained a M box of 61 amino acids in the N terminal.The secondary structure of CmFLC was mainly composed of alpha helix and random coil. CmFLC had a high similarity to other plant FLC proteins,and contained all the M box and K box.The three dimensional structure of CmFLC protein had high similarity with Populus tomentosa FLC.CmFLC and CcFLC were assigned to the same clade.Sense and antisense gene interference fragments were gained through specific primer amplification,size 320 bp according to enzyme digestion results.FLC interference vector was successfully constructed: pc1301-ubi-CMFLC-RNAi. Conclusion: homologous gene FLC existed in C.mollissima,and maybe participate in the regulation of flowering and vernalization.