国家自然科学基金(39800118)和霍英东青年教师基金(71030)资助项目.
选择青海高原海拔4000m高山上生长的唐古特红景天为实验材料,以叶片为外植体,在MS+BA2+NAA0.2固体培养基上诱导出黄绿色、松脆愈伤组织.愈伤组织细胞在同样成分的液体培养基中培养获得成功.在悬浮培养液中可检测到分泌蛋白的存在.经SDS-聚丙烯酰胺凝胶电泳分析表明,低温锻炼或脱落酸(ABA)诱导后,细胞分泌蛋白的多肽谱带数增加.与此相对应的是,细胞的抗冻能力也明显提高.PAS染色揭示多肽中均含有糖基.通过测定热滞值,确信细胞分泌蛋白是具有抗冻活性的糖蛋白.
The culture of callus and cell suspension induced from leaves of Rhodiola algida Var.tangutica collected from Qinghai plateau(altitude:4 000 m),on solid and in liquid culture medium(MS+BA2+NAA0.2)respectivelly,was successful.After cold (4~5℃)acclimation or ABA treatment for 10 or 14 days, the freezing resistance of suspension cells was increased. Proteins from the liquid suspension cell culture medium exhibited antifreeze activity. Treatment of the antifreeze proteins(AFPs) with boiling or trypsin caused inactivation,proving the protein characteristics of AFPs.Periodate and borate inhibited the antifreeze activity of AFPs,suggesting the presence of carbohydrate. Of the four treatments(25℃;25℃+ABA;5℃;5℃+ABA),2~3 polypeptides(29~50 ku) could be detected from the one dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE). PAS(periodic acid-schift’s reagent) reaction of SDS-PAGE showed the presence of carbohydrate in the polypeptides.
卢存福,简令成,匡廷云.低温诱导唐古特红景天细胞分泌抗冻蛋白[J].生物化学与生物物理进展,2000,27(5):555-559
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