橡胶树叶柄愈伤组织乳管细胞的组织化学和分子生物学研究
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摘要
橡胶树乳管分化研究主要以树皮为研究材料。本研究通过组织化学染色法、尼罗红荧光染色法、免疫组织化学法和分子生物学方法证实橡胶树叶柄来源的愈伤组织中存在乳管细胞。乳管细胞在叶柄愈伤组织中随机分布,分布模式类似橡胶树初生乳管;叶柄愈伤组织乳管细胞在发育早期时橡胶粒子较少,在发育后期时充满橡胶粒子。RT-PCR扩增出叶柄愈伤组织乳管细胞的橡胶延伸因子(REF)、小橡胶粒子蛋白(SRPP)、橡胶凝集素(hevein)和橡胶转移酶(CPT)等胶乳特异基因的转录本,经比对它们序列与所报道的橡胶树树干胶乳中表达的基因序列一致,表明橡胶树叶柄愈伤组织乳管细胞拥有树皮乳管细胞相似的功能,为叶柄愈伤组织作为一种新型的研究乳管分化的模式提供了科学依据。
The trunk bark of rubber tree(Hevea brasiliensis) is usually used as the material for the study of laticifer differentiation. In this study, we confirmed the presence of laticifer cells in petiole-derived calli by histochemical staining, nile red fluorescence staining, immunohistochemistry and molecular biology. Laticifer cells in petiole-derived calli were distributed randomly, similar to the primary laticifers. They contained less rubber particles in the early developmental stage, and were full of rubber particles in the late developmental stage. The laticifer-specific genes including REF, SRPP, Hevein and CPT were confirmed to be expressed from the petiole-derived callus by RT-PCR, and their transcripts were the same as those reported in the latex of the trunk barks. These results indicated that the laticifer cells in the petiole-derived calli and in barks had similar functions, which would provide a basis for petiole-derived callus as a novel research model to study laticifer differentiation in the rubber tree.
The trunk bark of rubber tree(Hevea brasiliensis) is usually used as the material for the study of laticifer differentiation. In this study, we confirmed the presence of laticifer cells in petiole-derived calli by histochemical staining, nile red fluorescence staining, immunohistochemistry and molecular biology. Laticifer cells in petiole-derived calli were distributed randomly, similar to the primary laticifers. They contained less rubber particles in the early developmental stage, and were full of rubber particles in the late developmental stage. The laticifer-specific genes including REF, SRPP, Hevein and CPT were confirmed to be expressed from the petiole-derived callus by RT-PCR, and their transcripts were the same as those reported in the latex of the trunk barks. These results indicated that the laticifer cells in the petiole-derived calli and in barks had similar functions, which would provide a basis for petiole-derived callus as a novel research model to study laticifer differentiation in the rubber tree.
引文
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[14]Tan D,Hu X,Fu L,et al.Comparative morphology and transcriptome analysis reveals distinct functions of primary and secondary laticifer cells in rubber tree[J].Scientific Reports,2017,7(1):3126.
[15]张治礼,郝秉中,吴继林.巴西橡胶树乳管分化的研究[J].热带作物研究,1996(4):4-10.
[16]范思伟.产胶生理[M].儋州:华南热带作物研究院,1983:33-34.
[17]谭德冠,吴煜,张家明.巴西橡胶树离体材料的绿色荧光背景研究[J].热带作物学报,2008,29(1):23-26.
[18]蔡太生.免疫组织化学技术[J].世界最新医学信息文摘,2004,3(6):1381.
[2]曾霞,黄华孙.国内外橡胶树种质资源收集保存及其研究进展[J].热带农业科技,2004,27(1):24-28.
[3]Gomez J B.Anatomy of Hevea and its influence on latex production[M]//Annual Report.Kuala Lumpur:Malaysia Rubber Research and Development Board,1982:76.
[4]Tan D,Sun X,Zhang J.Histochemical and immunohistochemical identification of laticifer cells in callus cultures derived from anthers of Hevea brasiliensis[J].Plant Cell Reports,2011,30(6):1117-1124.
[5]Tan D,Sun X,Zhang J.Age-dependent and jasmonic acid-induced differentiation of laticifer cells in anther callus cultures of rubber tree[J].Planta,2014,240(2):337-344.
[6]广东省农垦总局,海南省农垦总局.橡胶树良种选育与推广[M].广州:广东科技出版社,1994:20.
[7]田维敏,史敏晶,谭海燕,等.橡胶树树皮结构与发育[M].北京:科学出版社,2015:245-246.
[8]Sando T,Hayashi T,Takeda T,et al.Histochemical study of detailed laticifer structure and rubber biosynthesis-related protein localization in Hevea brasiliensis using spectral confocal laser scanning microscopy[J].Planta,2009,230(1):215-225.
[9]Tang C,Yang M,Fang Y,et al.The rubber tree genome reveals new insights into rubber production and species adaptation[J].Nature Plant,2016,2(6):1-10.
[10]吴华玲,段翠芳,聂智毅,等.巴西橡胶树橡胶粒子研究进展[J].热带作物学报,2009,30(7):1044-1049.
[11]Kwon M,Kwon E J,Ro D K.cis-prenyltransferase and polymer analysis from a natural rubber perspective[J].Methods Enzymology,2016(576):121-145.
[12]Berthelot T,Peruch F,Lecomte S.Highlights on Hevea brasiliensis(pro)hevein proteins[J].Biochimie,2016,127:258-270.
[13]史自强,胡正海.植物含橡胶组织的制片方法[J].植物学报,1965,13(2):179-182.
[14]Tan D,Hu X,Fu L,et al.Comparative morphology and transcriptome analysis reveals distinct functions of primary and secondary laticifer cells in rubber tree[J].Scientific Reports,2017,7(1):3126.
[15]张治礼,郝秉中,吴继林.巴西橡胶树乳管分化的研究[J].热带作物研究,1996(4):4-10.
[16]范思伟.产胶生理[M].儋州:华南热带作物研究院,1983:33-34.
[17]谭德冠,吴煜,张家明.巴西橡胶树离体材料的绿色荧光背景研究[J].热带作物学报,2008,29(1):23-26.
[18]蔡太生.免疫组织化学技术[J].世界最新医学信息文摘,2004,3(6):1381.