CAJ | 학술논문

从菊芋(Helianthus tuberosus L.)块茎cDNA文库中得到了一个新的植物类金属硫蛋白基因htMT2的cDNA序列,全长509 bp,包括240 bp的开放阅读框、62 bp的5′端非翻译区、207 bp的3′端非翻译区.通过PCR获得了2个htMT2编码区的部分基因组片段htMTG-1及htMTG-2,长度分别为986 bp和982 bp.分析表明两个基因组片段均包含3个外显子及2个内含子,编码一个由79个氨基酸残基组成的多肽,与从htMT2推测的多肽完全一致,该多肽具有植物类金属硫蛋白的典型结构特征,N端及C端结构域富含Cys,分别具有8个和7个Cys残基,上述两个结构域被一个无Cys的中间区分开.Southern杂交结果表明,htMT2在菊芋基因组中以小基因家族的形式存在.Northern杂交结果表明htMT2在叶片、叶柄、茎及块茎中均有表达,在茎中有较高水平的表达,但在根中未检测到杂交信号.经Cu2+处理后,htMT2在茎中的表达量显著降低.与其他2型金属硫蛋白的序列同源性比较及htMT2对金属离子处理的反应均表明,htMT2是一种新的植物类金属硫蛋白基因.
종국우(Helianthus tuberosus L.)괴경cDNA문고중득도료일개신적식물류금속류단백기인htMT2적cDNA서렬,전장509 bp,포괄240 bp적개방열독광、62 bp적5′단비번역구、207 bp적3′단비번역구.통과PCR획득료2개htMT2편마구적부분기인조편단htMTG-1급htMTG-2,장도분별위986 bp화982 bp.분석표명량개기인조편단균포함3개외현자급2개내함자,편마일개유79개안기산잔기조성적다태,여종htMT2추측적다태완전일치,해다태구유식물류금속류단백적전형결구특정,N단급C단결구역부함Cys,분별구유8개화7개Cys잔기,상술량개결구역피일개무Cys적중간구분개.Southern잡교결과표명,htMT2재국우기인조중이소기인가족적형식존재.Northern잡교결과표명htMT2재협편、협병、경급괴경중균유표체,재경중유교고수평적표체,단재근중미검측도잡교신호.경Cu2+처리후,htMT2재경중적표체량현저강저.여기타2형금속류단백적서렬동원성비교급htMT2대금속리자처리적반응균표명,htMT2시일충신적식물류금속류단백기인.
A novel cDNA sequence htMT2, which encodes a type 2 metallothionein-like protein, was isolated from Helianthus tuberosus L. tuber cDNA library. The whole sequence is 509 bp, including an open reading frame (ORF) of 240 bp, a 5′ UTR of 62 bp and a 3′ UTR of 207 bp. Two genomic sequences covering the coding region of htMT2 were cloned by PCR reaction. Sequence analysis revealed that the genomic sequences htMTG-1 of 986 bp and htMTG-2 of 982 bp were both composed of three exons and two introns. The deduced protein consisted of 79 amino acid residues with a predicted molecular weight of 7.8 ku (kD). Amino-terminal and carboxy-terminal domains contained 8 and 7 cysteine residues respectively, separated by a central cysteine free spacer. Sequence alignment revealed that the predicted protein of htMT2 was homologous to type 2 metallothioneins (MTs) of plants. Southern blotting analysis indicated that htMT2 was encoded by a small multi-gene family in H. tuberosus genome. Northern blotting analysis showed that htMT2 transcripts were detected in stems, leaves and leafstalks, but no transcripts were detected in roots. The expression level in stems was the highest among the above tissues. Transcripts in stems were significantly reduced by Cu2+ treatment. Judging from the homologies between the deduced HtMT2 and other type 2 plant metallothioneins as well as responses to metal ions, we believe that htMT2 encodes a new type 2 metallothionein.