摘要
【目的】探寻赤霉素诱导甘蔗节间伸长基因表达差异的分子基础,为分离节间伸长相关基因奠定基础。【方法】以新台糖22号为材料,在伸长初期以浓度为200mg·L-1的赤霉素进行叶面喷施,以喷施清水为对照,分别提取总RNA进行等量混合,获得2种不同处理的样品池,采用cDNA-SRAP技术进行基因差异表达分析。【结果】700对引物组合共扩增到约15000条、大小在50—750bp的cDNA片段。在GA3处理组和对照组中,甘蔗幼茎中的转录水平具有很大的差异,获得134条差异条带。从反向Northern杂交结果阳性的S-TDFs中选取24个片段进行克隆和序列分析,按照功能可以将24个差异表达基因片段S-TDFs(cDNA-SRAP transcriptderive dfragments)分为6类,分别为能量与代谢相关基因(4条,占总数的16.67%)、未知功能蛋白基因(6条,占总数的25%)、未知基因(10条,占总数的41.66%)、细胞壁生物合成与修饰相关基因(1条,占总数的4.17%)、信号传导相关基因(2条,占总数的8.33%)和转录因子相关基因(1条,占总数的4.17%)。【结论】cDNA-SRAP完全适用于差异表达分析,从中获得一些与甘蔗节间伸长相关的差异基因片段,这些基因可用于甘蔗节间伸长的分子机理研究。
Objective The research was carried out to explore the molecular basis of the differential expression of GA-induced stem elongation genes of sugarcane and isolation of elongation-related genes.Method The plants were sprayed with gibberellin（GA3） at concentration of 200 mg·L-1 at early elongation stage,with water spraying as control.After extraction,equal quantity of the total RNAs of the same treatment was mixed to get two different mixed RNA pools for the GA3 treatment and the control.By cDNA-SRAP technique,different expressions of the genes were analyzed.Result Using 700 pairs of primers,about 15 000 cDNA fragments in size between 50-750 bp were amplified.Some fragments were different between the GA3 treatment and the control for different gene transcriptions in the young stem of sugarcane.Among them,134 different cDNA fragments were selected.After reverse Northern blot analysis,24 positive S-TDFs（cDNA-SRAP transcript derived fragments） were selected for cloning and sequencing analysis.After BLAST analysis,the 24 S-TDFs were divided into six categories according to gene function：energy and metabolism-related genes（4,accounting for 16.67%）,genes of unknown functional proteins（6,accounting for 25%）,unknown genes（10,accounting for 41.66%）,cell wall biosynthesis and modification related genes（2,accounting for 8.33%）,and transcription factor-related genes（1,accounting for 4.17%）.Conclusion This study shows that the cDNA-SRAP technique is applicable to the analysis of gene differential expression.Some fragments of differentially expressed genes associated with sugarcane stem elongation were obtained,and may be used for investigating the molecular mechanism of sugarcane stem elongation.
出处
《中国农业科学》
CAS
CSCD
北大核心
2010年第19期3937-3944,共8页
Scientia Agricultura Sinica
基金
国家"十一五"科技支撑计划项目(2007BAD30B03)
广西科技攻关重点项目(桂科攻0782004-3)
广西科技攻关项目(桂科能0630006-5F
桂科能0815011)
现代农业产业技术体系建设专项资金(nycytx-024-01-15)
