Identification and characterization of genes expressed in early embryogenesis from microspores of Brassica napus
Identification and characterization of genes expressed in early embryogenesis from microspores of Brassica napus
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DOI：
10.1007/s00425-006-0388-8
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发表时间：
2007-02
期刊：
影响因子：
4.3
通讯作者：
Ryo Tsuwamoto;H. Fukuoka;Y. Takahata
中科院分区：
文献类型：
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作者：
Ryo Tsuwamoto;H. Fukuoka;Y. Takahata
为了了解甘蓝型油菜小孢子诱导胚胎发生的机制，我们对小孢子培养早期表达差异的基因进行了详尽的分离。通过抑制减法杂交得到雄激素启动过程中上调基因的减法cDNA文库，并用斑点杂交进行差异筛选，共鉴定出136个非冗余表达序列标签。基因潜在功能分析表明，其中64%的基因与已知基因同源，其余基因未见参与胚胎发生的报道。分离的基因中含有许多胚胎特异性基因，如脂质转移蛋白、napin、十字花素、油蛋白、植物磺酸等基因。对15个已知与胚胎发生无关的基因进行实时RT-PCR分析，发现所有基因在小孢子胚胎发生早期都有高表达。少数基因在非胚性条件下或胚胎后期培养的小孢子中也有较高的表达。对15个基因的表达谱进行主成分分析，结果表明，这些基因可分为两类，一类在胚胎发生初期高表达，另一类在胚胎发生早期至中期高表达。这些基因的表达在合子胚胎中得到证实。本研究分离的基因的鉴定和特性为油菜小孢子胚胎发生提供了新的信息。
To understand the mechanism in induction of embryogenesis from microspores ofBrassica napus, we isolated exhaustively the genes expressed differentially during the early stage of microspore culture. A subtracted cDNA library composed of up-regulated genes during androgenic initiation was produced by suppression subtractive hybridization followed by differential screening by dot blot hybridization, and a total of 136 non-redundant expressed sequence tags were identified. Analysis of the potential functions of the genes showed that 64% of these genes were homologous to known genes, and the remaining ones have not been previously reported to participate in embryogenesis. Many embryo-specific genes were contained in the isolated genes, for example, genes cording lipid transfer protein, napin, cruciferin, oleosin, and phytosulfokine. Real-time RT-PCR analysis for 15 selected genes, which are understood to not be related with embryogenesis, demonstrated that all genes were expressed highly in the early stage of microspore embryogenesis. A few genes also showed higher expression in microspores cultured in non-embryogenic condition or in later stages of embryos. A principal component analysis based on expression profiles of the 15 genes demonstrated that these genes were classified into 2 groups, one characterized by their high expression in initiation of embryogenesis, and the other characterized by their expression in the early to middle stage of embryogenesis. The expressions of these genes were confirmed in zygotic embryos. The identification and characterization of the genes isolated in the present study provide novel information on microspore embryogenesis inBrassica.