尾叶桉不同类型愈伤组织APX类基因转录水平及酶活性的差异

Differences of APX-genes Transcription Level and Enzyme Activity in Different Types of Eucalyptus urophylla Callus

比较尾叶桉(Eucalyptus urophylla)不同类型愈伤组织抗坏血酸过氧化物酶(APX)酶活性及APX基因转录水平,探讨愈伤分化与不同位置APX表达的关系。将下胚轴接种在愈伤诱导培养基中,外植体膨大后先形成白色愈伤组织,随后继续膨大并分化成白色、红色、绿色愈伤组织,绿色愈伤组织可进一步分化形成带芽点绿色愈伤组织。检测不同愈伤组织APX酶活性,qPCR比较过氧化物酶体(APX1-3)、细胞质(APX4-6)和叶绿体(APX7-9)中共9个APX基因的转录水平。4种不同类型愈伤组织的APX酶活性由大到小依次为红色、绿色、带芽点绿色和白色愈伤组织。与红色愈伤组织相比,绿色愈伤组织和带芽点绿色愈伤组织中的APX1、APX2、APX3、APX5、APX6、APX7、APX8的转录水平均增强。设红色愈伤组织基因的相对表达量为1,这7个基因在绿色愈伤组织中的相对表达水平分别为1.416、1.917、1.612、2.719、2.321、1.591、21.004;7个基因在带芽点绿色愈伤组织中的相对表达水平分别为2.313、2.517、2.612、3.856、1.493、2.534、29.357。除APX6外,其余6个基因转录水平均为带芽点绿色愈伤组织最高,红色愈伤组织最低。结果表明,过氧化物酶体、细胞质和叶绿体中的APX基因总体表达上调促进了芽原基的分化。绿色愈伤和带芽点绿色愈伤中叶绿体APX8转录水平显著升高,说明APX8表达与叶绿体发育密切相关。

In order to investigate the relationship between callus differentiation and APX-genes expressionin Eucalyptus urophylla,the ascorbate peroxidase(APX)activity and the APX transcription level of different types callus were evaluated.Inoculated on the SPCa medium,the hypocotyl explants formed white callus.Then,the white callid differentiated into three types,including white calli,red ones,and green ones.The green ones further differentiated into green buds.These calli were used to evaluate the APX activities and the relative transcription levels of nine APX genes.Three APX genes located in peroxisome were numbered as APX1,APX2,APX3,three cytoplasmic APX genes were numbered as APX4,APX5,APX6,and three chloroplast APX genes were numbered as APX7,APX8,APX9.The APX activities of four different types of calli were red calli,green ones,green ones with buds and white ones from high to low.Compared with red calli,the transcription levels of APX1,APX2,APX3,APX5,APX6,APX7 and APX8 in green calli and green calli with buds were enhanced.By normalizing the transcription levels of corresponding genes in red callus to 1,the transcription levels of the seven genes in green calli were 1.416,1.917,1.612,2.719,2.321,1.591 and 21.004,respectively.The transcription levels of these seven genes in green calli with buds were 2.313,2.517,2.612,3.856,1.493,2.534 and 29.357,respectively.Except for APX6,the transcription levels of the other 6 genes in green calli with buds were the highest and those in red calli were the lowest.The up-regulation of APX gene in peroxisome,cytoplasm and chloroplast promoted the differentiation of bud primordia.The chloroplast APX8 increased significantly in green calli and green ones with buds implied that the expression of APX8 was closely related to chloroplast development.The study might provide reference for further research on the molecular mechanism of Eucalyptus callus differentiation.