CAJ | 학술논문

Wap25是从桑树幼茎克隆的低温诱导蛋白基因,为胚胎后期富集蛋白(LEA)基因的成员之一,与其它物种低温诱导基因的同源性很低.生物信息学分析表明,桑树低温诱导基因Wap25编码蛋白由226个氨基酸组成,分子质量25.3 kD,等电点5.52;蛋白N端1-30位氨基酸表现出强烈的疏水性,其后是亲水性高的区域,平均亲水值达-1.020,而蛋白1-29位氨基酸为典型的真核生物蛋白质的信号肽序列,表明WAP25蛋白属于分泌型蛋白.用SubLocv 1.0软件预测WAP25的亚细胞定位,将该蛋白定位于细胞质中.Tmpred软件预测WAP25不存在跨膜结构.构建Wap25与绿色荧光蛋白(GFP)的重组质粒,用基因枪法将其转入洋葱表皮细胞,在激光扫描共聚焦显微镜下发现GFP分散于洋葱表皮的细胞质中,而在细胞核未发现GFP的表达,此结果与WAP25的亚细胞定位预测结果相符.
Wap25시종상수유경극륭적저온유도단백기인,위배태후기부집단백(LEA)기인적성원지일,여기타물충저온유도기인적동원성흔저.생물신식학분석표명,상수저온유도기인Wap25편마단백유226개안기산조성,분자질량25.3 kD,등전점5.52;단백N단1-30위안기산표현출강렬적소수성,기후시친수성고적구역,평균친수치체-1.020,이단백1-29위안기산위전형적진핵생물단백질적신호태서렬,표명WAP25단백속우분비형단백.용SubLocv 1.0연건예측WAP25적아세포정위,장해단백정위우세포질중.Tmpred연건예측WAP25불존재과막결구.구건Wap25여록색형광단백(GFP)적중조질립,용기인창법장기전입양총표피세포,재격광소묘공취초현미경하발현GFP분산우양총표피적세포질중,이재세포핵미발현GFP적표체,차결과여WAP25적아세포정위예측결과상부.
Wap25 gene,which belongs to the late embryogenesis abundant(LEA)family,was cloned from young stem of mulberry.The alignment analysis showed that this gene had very low identities with cold-induced genes in other spe-cies.Bioinformatics analyses indicated that Wap25 encodes a 226 amino acid protein with a theoreticaI molecular mass of 25.3 kD and a pl of 5.52.Its N-terminal amino acids 1 to 30 have strong hydrophobicity.The following amino acids are highly hydrophilic,with an average hrdrophilicity score of-1.020.The N-terminus contains a typical signal peptide of eukaryotic proteins with a putative cleavage site located after position 29,suggesting that WAP25 is a secreted protein.Subcellular localization prediction with SubLocv 1.0 program indicated that WAP25 was Iocated in cytoplasm and predic-tion with Tmpred software revealed that WAP25 had no transmembrane domain.A green fluorescent protein(GFP)fu-sion recombinant protein expression vector was constructed and transformed into onion epidermal cells by biolistic meth-od.Observation under laser scanning confocal microscope showed that GFP was disseminated in the cytoplasm of onion epidermal cells and no signal was found in nucleus,being consistent with the predicted result of subcellular localization,