大竹蛏水管自切后再生过程的转录组学分析

Transcriptome analysis of regenerative process after autotomy of siphon in jackknife clam(Solen grandis)

为探究大竹蛏(Solen grandis)水管自切后再生期间的转录组变化情况,通过二代测序技术分别对大竹蛏(壳长为8~10 cm)水管自切前(对照组)和自切后7 h、7 d、20 d(选择组)进行转录组学分析。结果表明:原始数据经拼接注释后共获得103909条Unigenes,其中,30027条Unigenes得到注释,注释率为28.90%;差异基因GO功能注释显示,自切后7 h时钙离子结合条目富集程度最高且富集基因数最多,自切后7 d和20 d时几丁质结合条目富集程度最高;KEGG通路富集分析显示,仅自切后7 d时具有显著富集通路,包括氨基糖与核苷酸糖代谢、氮素代谢、黏着斑、溶酶体和MAPK信号通路;差异基因时间趋势分析显示,富集基因数前3的模式依次为Profile5、Profile17和Profile12,各模式表达基因GO功能注释结果显示,Profile5中有529条基因显著富集到离子结合条目,占比为53.77%,Profile17和Profile12中显著富集基因数最多的条目分别为受体活性和几丁质代谢;随机挑选6个差异基因进行RT-qPCR验证,结果证明转录组测序结果可靠。初步推测,在水管再生早期(自切后7 h),以钙离子为代表的信号分子在传递受损信号过程中发挥重要作用,同时多种受体活性增强响应水管再生过程,随着再生的进行,物质代谢和生物合成过程大量增加以满足组织再生的需求。

In order to explore the changes in transcriptome of jackknife clam(Solen grandis)after siphon autotomy,the transcriptome of jackknife clam with shell length of 8-10 cm before(control group)and 7 h,7 d and 20 d after siphon autotomy(selection group)were analyzed by second-generation sequencing technology.The results showed that 103909 Unigenes were obtained from the raw data after splicing and annotation,in which 30027 Unigenes were annotated,with an annotation rate of 28.90%.The gene ontology functional enrichment results of three groups of differentially expressed genes showed that the calcium-binding items were the most enriched at 7 hours after autotomy,and the chitin-binding items were the most enriched at 7 days and 20 days after autotomy.The results of KEGG enrichment pathway showed that the jackknife clam had significantly enriched pathways only 7 days after autotomy,including amino sugar and nucleotide sugar metabolism,nitrogen metabolism,focal adhesion,lysosome and MAPK signal pathway.The time trend analysis of all differentially expressed genes in three groups showed that the top three profiles of gene enrichment were Profile5,Profile17 and Profile12.The gene ontology function annotation results of the three patterns showed that 529 genes in Profile5 were significantly enriched to ion-binding items,accounting for 53.77%,and the items with the largest number of genes enriched were receptor activity in Profile17 and chitin metabolic activity in Profile12.The RT-qPCR verification of six differentially expressed genes proved that the transcriptome sequencing results were reliable,and that preliminarily speculated various signal molecules represented by calcium ions played an important role in the process of transmitting damaged signals in the early stage of siphon regeneration(7 hours after autotomy),meanwhile the activity of various receptors was enhanced in response to the siphon regeneration process.With the progress of regeneration,the processes of substance metabolism and biosynthesis were greatly increased to meet the needs of tissue regeneration.