龙须菜不同世代间SSH文库差显基因的信息学分析及部分序列的转录验证

Bio-Informatic Analysis of the Differential Expressed Sequences from SSH Libraries Between Different Phases of Gracilariopsis lemaneiformis and the Transcriptive Validation for Several Candidate Genes

本研究通过对5个龙须菜(Gracilariopsis lemaneiformis)抑制性消减杂交文库(SSH文库)的1 873条差异显示序列进行聚类和生物信息学分析,共获得118个重叠群以及247条未被聚类进任何重叠群的单条序列。Blast2GO注释分析结果说明龙须菜性别和世代的遗传是由多因子参与、多细胞共同完成的复杂生命活动。选取8条差显序列进行了实时荧光定量PCR分析以验证数据的可靠性,发现尽管SSH文库具有较高的假阳性率,但对斑点杂交结果进行进一步的聚类分析和RT-qPCR验证后可以有效提高实验结果的可信度,并从中发现contig03和7310很可能真正与雌配子体世代相关。

Through clustering and then bioinformatics analysis for those 1 873 differential displayed sequences acquired from 5 pairs of suppression subtractive hybridization(SSH)libraries of the Gracilariopsis lemaneiformis established in previous study,totally 118 contigs and 247 single sequences which weren’t clustered into any contig were obtained in this study.The results of blast and annotation by Blast2GO confirmed the conclusion that various affecting factors and multiple cellular functions were involved in the genetic processes related to sex or phases change in G.lemaneiformis.Furthermore,8 candidate sequences were selected to analyze the reliability of SSH data using Real-time Quantitative PCR.Although the false positive rate was quite high for SSH,further cluster analysis and RT-qPCR verification of dot-blot results could effectively improve the reliability of the experimental results.And contig03 and single sequence 7310 were proved with high probability related to the phases of female gametophyte.