一个新的Trihelix转录因子家族基因在油菜抗逆应答和种子发育中的功能研究

Along with the gradually aggravating of soil drought and salinization problems, China's edible oil gap is worsening. Creating drought- and salt-tolerant, yield-increasing rapeseed varieties is very important and urgent for the protection of national edible oil supply security. On the basis of preliminary studies, we discovered a new gene from Brassica napus with the potential functions of abiotic-stress tolerance and yield-enhancement. This gene is homologous to the SIP1 subfamily genes of Trihelix transcription factor family of Arabidopsis, and it is the first gene of such family identified in Brassica napus, so we named it BnSIP1-1. We found that the tolerance of the transgenic Arabidopsis over-expressing BnSIP1-1 to abiotic stress (Mannitol, Nacl and ABA) was improved, seed size was bigger and thousand-seeds weight increased. The project intends to study the functions of BnSIP1-1 in abiotic-stress tolerance and roles in seed developmental process in Brassica napus by reverse genetics methods. The function mechanism will be dissected from phenotype to cytological level, physiological and biochemical level, and metabolism level. The use of digital gene expression profiles technique will preliminary clarify the gene regulation networks involved in stress resistance and seed development. This project will not only fill the blank of functional research of the Trihelix family gene in Rapeseed, and deepen the understanding of the theory of stress response mechanisms and seed development mechanism of Rapeseed, but also provide a theoretical basis to foster stress-resistant, high-yield new rapeseed varieties.

土壤干旱和盐碱化问题的愈演愈烈使得我国食用油缺口日趋严重，培育抗旱耐盐、增产稳产油菜品种对于保障我国食用油供给安全十分重要和紧迫。申请者在前期研究基础上从油菜中挖掘到一个可能同时具有抗逆和增产功能的新基因，该基因与拟南芥Trihelix转录因子家族SIP1亚家族基因同源，是目前第一个在油菜中被鉴定的该家族成员，命名为BnSIP1-1。在拟南芥中过表达该基因不仅提高植株抗逆（甘露醇、Nacl和ABA）能力，而且引起种子变大千粒重增加。本项目拟通过反向遗传学法在油菜中超表达和抑制该基因，由表型深入到细胞、生理生化代谢水平确认该基因在逆境胁迫应答和种子发育过程中的功能，利用数字基因表达谱技术初步分析该基因参与抗逆和种子发育的分子机制。本项目的研究不仅会填补油菜中Trihelix家族基因功能研究的空白，而且将深化对油菜胁迫应答机制和种子发育机理的理论认识，为培育抗逆、高产的油菜新品种提供理论基础。

研究与抗逆、增产相关的基因及其作用机制对于保证粮食安全具有重要的理论意义和应用价值。申请者在前期研究的基础上从油菜中挖掘到一个可能同时具有抗逆和增产功能的新基因，该基因与拟南芥Trihelix转录因子基因家族中SIP1亚家族成员同源，是目前第一个在油菜中被鉴定的该家族基因，命名为BnSIP1-1。我们对BnSIP1-1基因的基本生物学特征进行了分析，发现其编码的蛋白质C端含有Trihelix转录因子基因家族的保守结构域trihelix，N端含有一个保守的helix结构域；对该基因启动子功能元件的预测发现它含有多个光应答元件、脱落酸响应元件（ABRE motif）、胁迫应答元件（TC-rich motif），以及与热激和低温应答相关的HSE和LTR元件；蛋白定位于细胞核；是一个泛组织表达的基因；该基因的转录水平会随着外施的水分胁迫、盐胁迫和ABA胁迫的浓度以及胁迫时间发生改变。在拟南芥中过表达BnSIP1-1会提高植株对甘露醇和ABA的耐受性，另外还导致转基因种子变大。在油菜中过表达BnSIP1-1同样会提高植株对甘露醇和ABA的耐受性。经甘露醇处理后，转基因材料叶片的叶绿素含量明显高于野生型；经ABA处理，转基因材料的发芽率和幼苗期的根长、茎长、根重以及地上部分净重明显高于野生型。对BnSIP1-1参与的调控网络进行了初步分析，BnSIP1-1主要参与调节ABA通路及水分胁迫相关基因的表达，如BnRD29A、BnERD15、BnLEA1和BnNAC485，而与盐胁迫早期应答相关的基因BnSOS1和 BnNHX1的表达水平并未受到BnSIP1-1的影响。通过本项目对BnSIP1-1功能的系统剖析，我们明确了该基因在植物逆境胁迫应答过程中的功能，对其参与的调控网络进行了分析，为深入探索油菜逆境响应机制提供了重要的理论支撑。