【摘要】 用RT-PCR和RACE-PCR技术,从热激处理的甜椒叶片总RNA中扩增出了细胞质小分子量热激蛋白(sHSP)全长779bp的cDNA基因序列,包含一个480bp开放阅读框,编码159个氨基酸。Southern杂交结果表明在甜椒基因组中有该基因的小的多基因家族。Northern结果显示该基因在甜椒根、茎、叶中的表达受热激和低温的诱导。原核表达分析表明该基因在高温以及低温条件下可以提高大肠杆菌的生存能力。更多还原

【Abstract】 Full length 779 bp cytosolic clsss IsHSP cDNA was cloned from heat-shockedsweet pepper leaves using a PCR approach withdegenerate primers designed from conservedmotifs found in a number of plant cytosolic clsss IsHSP genes, named CaHSP18. Its accessionnumber is AY284925. CaHSP18 was high iden-tified with Nicotiana tabacum (Fig.1). A multi-gene family was found in sweet pepper genomicDNA by Southern-blot analysis (Fig.3). North-ern blot revealed that the expression ofCaHSP18 in sweet pepper was induced by heattreatment and was significantly different be-tween different tissues (Fig.4). The transcrip-tion of CaHSP18 in leaves and stems werehigher than those in roots. The expression ofCaHSP18 could be detected after chilling treat-ment at 4℃ for 2 d. Recombinant CaHSP18was overexpressed in Escherichia coli to studyits possible function under heat and chillingstress. Sodium dodecyl sulfate-polyacrylamidegel electrophoresis analysis of cell lysates sug-gested that CaHSP18 was expressed in Escheri- chia coli (Fig.6). The growth of wild type andtransformed cells was similar at 37℃ (Fig.5).Upon transfer from 37℃ to 50℃, a temperatureknown to cause cell autolysis, those cells thataccumulated CaHSP18 showed improved viabil-ity compared with the control lines (Fig.7). Totest the hypothesis that sHSPs may be involvedin protection against chilling stress, the viabilityof recombinant cells at 4℃ was studied. Thesweet pepper CaHSP18 significantly enhancedcell survivability (Fig.8). These results indicatethat plant cytosolic clsss I sHSP have protectivefunctions not only against heat stress but alsoagainst chilling stress.更多还原