光诱导的马铃薯试管薯形成相关基因转录组分析
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摘要
马铃薯(Solanum tuberosum L.)作为世界第四大粮食作物,在世界粮食安全中发挥着重要作用。块茎是马铃薯植株的地下变态茎,由匍匐茎在条件适合时停止纵向伸长并在匍匐茎亚顶端膨大形成,是主要经济器官。同时,马铃薯块茎还是研究植物变态器官发育的模式体系。因此马铃薯块茎发育及其调控的分子机理不但具有重要的经济意义,还有突出的生物学意义。光照特别是光质和光周期是影响马铃薯块茎形成的重要因素。目前,光质对组织培养条件下块茎形成影响的研究还不系统；光周期调控马铃薯块茎形成是一个复杂的生物学过程,需要数量众多的基因以及各种通路共同协作才能完成,尽管相关研究已取得了一定进展,但其分子机理还没有系统证实。新的基因组学研究技术和工具的出现,特别是高通量测序技术的出现将对分离和鉴定参与块茎形成过程的关键基因产生极大的推动作用。
本研究的主要目的是研究光质对组织培养条件下马铃薯试管薯形成的影响,进而为马铃薯试管薯生产中的光源选择提供理论支持；分离参与光周期调控马铃薯试管块茎形成的关键候选基因,为光周期调控马铃薯试管块茎形成分子机理的研究提供新的基因资源。获得的主要结果如下：
1)系统研究了白、红、蓝三种光质对组培养条件下马铃薯试管薯形成的影响,通过对单株结薯数、单薯重等的观察,表明基础苗繁殖光质、试管薯诱导期光质以及基因型对试管薯形成有显著影响,且各因素间存在显著的交互作用。总体而言,组织培养条件下白光更利于马铃薯试管薯的形成。当完成试管块茎形态建成后,蓝光光源更利于试管薯的持续膨大。红光对组织培养下试管薯的形态建成及生长具有阻碍作用。
2)评价鉴定了E26、E20、E109等来自于同一个杂交组合的马铃薯姊妹系在组织培养条件下试管薯形成的能力,其中E26对光周期敏感,仅能在8h/d光照条件下形成块茎；E109对光周期不敏感,在8h/d和16h/d条件下均能形成块茎；E20对光周期不敏感,但在任一光照条件下均不能形成块茎。结果反映这三个具有相同遗传背景的品系是研究光周期调控块茎形成的理想材料,适于本研究。
3)采用Digital Gene Expression profiling (DGE)高通量测序技术分析光周期敏感型品系E26在诱导条件(短日照8h/d,SD)和非诱导条件(长日照16h/d, LD)下基因表达在转录水平上的差异,共分离得到了2218个差异表达基因(differentially expressed gene, DEG)。
4)对DEGs进行了Gene ontology (GO)和KEGG pathway功能注释,发现差异表达基因涉及117个代谢通路和21个生物学过程,说明光周期调控马铃薯试管块茎形成是一个复杂的生理过程,需要各类基因、不同代谢途径的协同调节。
5)根据GO和KEGG pathway注释,选择了56个涉及植物节律、信号传导、发育以及若干功能未知基因,通过比较其在三个来自于同一杂交组合但结薯习性对光周期响应不同的姊妹系中的表达模式,证明PGSC0003DMT400083080(DNAbinding with one finger domain class转录因子)、PGSC0003DMT400068851(蓝光受体)、PGSC0003DMT400034146(植物凝集素)、PGSC0003DMT400043211(syntaxin-like蛋白)以及一个未知功能基因(PGSC0003DMT400022042)可能参与了光周期对马铃薯试管薯形成的调控。
本研究为马铃薯试管薯生产的光源选择提供了理论支持；分离得到的参与光周期调控马铃薯试管薯形成的候选基因,为研究光周期调控马铃薯试管薯形成的分子机理提供了新的策略。
Potato(Solarium tuberosum L.), a member of the Solanaceae, ranks the fourth most important food crop after rice, wheat, and maize (http://www.fao.org/)., and plays a crucial role in global food security. Potato tubers—the modified underground stems that are rich in starch, protein, antioxidants and vitamins—are the most important organ of the species. Tuber formation has thus attracted considerable attention, with a view to improve yields and provide a model system to look into the development of modified organs in plants. Potato tuberization is influenced by light quality and photoperiod. Effects of light quality on potato tuber formation in vitro is not well studied. Photoperiodic regulation of potato tuber formation is a very complex biological process that requires the interaction of diverse genes and the crosstalk of several pathways. Photoperiodic control of potato tuber formation is far from been fully elucidated. New genomics tools, especially high-throughput sequencing, should considerably facilitate the identification of candidate genes involved in the photoperiodic control of tuber formation in potato.
The main purposes of present research are to investigate the effects of light quality on potato tuberization in vitro for efficient selection of light source for microtuber production, and to identify candidate genes involved in the photoperiodic tuberization approaching insight the mechanism of photoperiodic regulation of potato tuberization. The main results obtained are as below.
1) By treating8potato genotyes with white, red and blue light in vitro, number of microtuber formed per plantlet and mean weight of the microtubers were investigated. The data showed that light quality for plant growth (LP), light quality for microtuberinitiation and growth (LM) and potato genotype impacted tuber formation dramatically. Generally, white light promoted microtuber formation while red light repressed the tuberization. Blue light was more suitable for tuber growth after tuber initiation.
2) Phenotypic characterization of E26, E20and E109, the three sister lines derived from same cross, indicated thattuberization of E26is photoperiod-sensitive characterized by tuber formation only occurs at short day (8h/d). E109is phtoperiod-independent for tuberization since it can form tubers at either short day or long day (16h/d), whereas E20is not capable to produce tubers at either of the daylength which is also considered photoperiod-independent. These three clones are desirable for research on the photoperiodic regulation of potato tuberization in present research.
3) Digital Gene Expression (DGE) Tag Profiling analysis of the short-day-sensitive clone E26identified2,218genes that were differentially regulated by day length.
4) Gene ontology (GO) and KEGG pathway analyses of DEGs found that DEGs involved117pathways and21biological processes. These results indicated that the response of potato to photoperiod requires the interaction of diverse genes and the crosstalk of several pathways.
5) According to GO and KEGG pathway annotation, we selected56genes associated with circadian rhythmicity, signal transduction, and development. Quantitative transcriptional analysis in the selected clones revealed five genes potentially associated with photoperiodic tuberization, which were predicted to encode a DOF protein (PGSC0003DMT400083080), a blue light receptor (PGSC0003DMT400068851), a lectin (PGSC0003DMT400034146), a syntaxin-like protein (PGSC0003DMT400043211), and a protein with unknown function (PGSC0003DMT400022042).
Our research provide a theoretical base for light source selection in microtuber production. Identification of the key candidate genes involved in photoperiodic-regulated tuberization gains insight the novel gene resources for further dissecting photoperiodic regulation of potato tuber formation in vitro.
本研究的主要目的是研究光质对组织培养条件下马铃薯试管薯形成的影响,进而为马铃薯试管薯生产中的光源选择提供理论支持；分离参与光周期调控马铃薯试管块茎形成的关键候选基因,为光周期调控马铃薯试管块茎形成分子机理的研究提供新的基因资源。获得的主要结果如下：
1)系统研究了白、红、蓝三种光质对组培养条件下马铃薯试管薯形成的影响,通过对单株结薯数、单薯重等的观察,表明基础苗繁殖光质、试管薯诱导期光质以及基因型对试管薯形成有显著影响,且各因素间存在显著的交互作用。总体而言,组织培养条件下白光更利于马铃薯试管薯的形成。当完成试管块茎形态建成后,蓝光光源更利于试管薯的持续膨大。红光对组织培养下试管薯的形态建成及生长具有阻碍作用。
2)评价鉴定了E26、E20、E109等来自于同一个杂交组合的马铃薯姊妹系在组织培养条件下试管薯形成的能力,其中E26对光周期敏感,仅能在8h/d光照条件下形成块茎；E109对光周期不敏感,在8h/d和16h/d条件下均能形成块茎；E20对光周期不敏感,但在任一光照条件下均不能形成块茎。结果反映这三个具有相同遗传背景的品系是研究光周期调控块茎形成的理想材料,适于本研究。
3)采用Digital Gene Expression profiling (DGE)高通量测序技术分析光周期敏感型品系E26在诱导条件(短日照8h/d,SD)和非诱导条件(长日照16h/d, LD)下基因表达在转录水平上的差异,共分离得到了2218个差异表达基因(differentially expressed gene, DEG)。
4)对DEGs进行了Gene ontology (GO)和KEGG pathway功能注释,发现差异表达基因涉及117个代谢通路和21个生物学过程,说明光周期调控马铃薯试管块茎形成是一个复杂的生理过程,需要各类基因、不同代谢途径的协同调节。
5)根据GO和KEGG pathway注释,选择了56个涉及植物节律、信号传导、发育以及若干功能未知基因,通过比较其在三个来自于同一杂交组合但结薯习性对光周期响应不同的姊妹系中的表达模式,证明PGSC0003DMT400083080(DNAbinding with one finger domain class转录因子)、PGSC0003DMT400068851(蓝光受体)、PGSC0003DMT400034146(植物凝集素)、PGSC0003DMT400043211(syntaxin-like蛋白)以及一个未知功能基因(PGSC0003DMT400022042)可能参与了光周期对马铃薯试管薯形成的调控。
本研究为马铃薯试管薯生产的光源选择提供了理论支持；分离得到的参与光周期调控马铃薯试管薯形成的候选基因,为研究光周期调控马铃薯试管薯形成的分子机理提供了新的策略。
Potato(Solarium tuberosum L.), a member of the Solanaceae, ranks the fourth most important food crop after rice, wheat, and maize (http://www.fao.org/)., and plays a crucial role in global food security. Potato tubers—the modified underground stems that are rich in starch, protein, antioxidants and vitamins—are the most important organ of the species. Tuber formation has thus attracted considerable attention, with a view to improve yields and provide a model system to look into the development of modified organs in plants. Potato tuberization is influenced by light quality and photoperiod. Effects of light quality on potato tuber formation in vitro is not well studied. Photoperiodic regulation of potato tuber formation is a very complex biological process that requires the interaction of diverse genes and the crosstalk of several pathways. Photoperiodic control of potato tuber formation is far from been fully elucidated. New genomics tools, especially high-throughput sequencing, should considerably facilitate the identification of candidate genes involved in the photoperiodic control of tuber formation in potato.
The main purposes of present research are to investigate the effects of light quality on potato tuberization in vitro for efficient selection of light source for microtuber production, and to identify candidate genes involved in the photoperiodic tuberization approaching insight the mechanism of photoperiodic regulation of potato tuberization. The main results obtained are as below.
1) By treating8potato genotyes with white, red and blue light in vitro, number of microtuber formed per plantlet and mean weight of the microtubers were investigated. The data showed that light quality for plant growth (LP), light quality for microtuberinitiation and growth (LM) and potato genotype impacted tuber formation dramatically. Generally, white light promoted microtuber formation while red light repressed the tuberization. Blue light was more suitable for tuber growth after tuber initiation.
2) Phenotypic characterization of E26, E20and E109, the three sister lines derived from same cross, indicated thattuberization of E26is photoperiod-sensitive characterized by tuber formation only occurs at short day (8h/d). E109is phtoperiod-independent for tuberization since it can form tubers at either short day or long day (16h/d), whereas E20is not capable to produce tubers at either of the daylength which is also considered photoperiod-independent. These three clones are desirable for research on the photoperiodic regulation of potato tuberization in present research.
3) Digital Gene Expression (DGE) Tag Profiling analysis of the short-day-sensitive clone E26identified2,218genes that were differentially regulated by day length.
4) Gene ontology (GO) and KEGG pathway analyses of DEGs found that DEGs involved117pathways and21biological processes. These results indicated that the response of potato to photoperiod requires the interaction of diverse genes and the crosstalk of several pathways.
5) According to GO and KEGG pathway annotation, we selected56genes associated with circadian rhythmicity, signal transduction, and development. Quantitative transcriptional analysis in the selected clones revealed five genes potentially associated with photoperiodic tuberization, which were predicted to encode a DOF protein (PGSC0003DMT400083080), a blue light receptor (PGSC0003DMT400068851), a lectin (PGSC0003DMT400034146), a syntaxin-like protein (PGSC0003DMT400043211), and a protein with unknown function (PGSC0003DMT400022042).
Our research provide a theoretical base for light source selection in microtuber production. Identification of the key candidate genes involved in photoperiodic-regulated tuberization gains insight the novel gene resources for further dissecting photoperiodic regulation of potato tuber formation in vitro.
引文
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