应用shell脚本批量分析肠道病毒Sanger测序结果

Application of shell script in batch processing for Sanger sequencing results of enteroviruses

目的编写shell脚本,批量完成柯萨奇病毒等肠道病毒Sanger测序结果的数据处理和比对分析,并为后续工作准备数据文件。方法选择已完成手工分析的柯萨奇病毒等肠道病毒VP1基因测序文件,编写shell脚本模拟手工分析程序,顺序使用phred、phd2fasta和phrap等三个软件和单机版NCBI-BLAST软件可完成从碱基识别至序列比对的全部工作。比对结束后使用Linux命令筛选信息,生成样本和匹配的序列名称文件、基因型以及用于后续分析的序列文件。使用MEGA 6.0软件比较手工分析得到的序列与shell脚本计算得到的序列之间的相似性,评价shell脚本分析的可靠性。结果使用shell脚本可完成所有功能。Shell脚本计算和手工分析的比对结果完全符合。结论shell脚本分析缩短了测序结果分析中重复工作耗费的时间,使研究人员可更加专注于序列后续分析。

Objective To create a shell script for the batch processing of Sanger sequencing results and sequence alignments of enteroviruses, e.g. coxsackievirus, and to prepared data files with standard format for further analysis. Methods The VP1 genes of enteroviruses, e.g. coxsackievirus were sequenced and the results were manually analyzed. A shell script was written to simulate manual analysis of the same sequence trace files. In the script, three softwares, phred, phd2fasta and phrap, and stand-alone NCBI-BLAST software were called in-turn to accomplish base-calling, file formatting, sequence assembly and alignment analysis. After alignment, a series of Linux text processing commands were used to generate files containing sample names, descriptions of reference sequences with the highest similarity, genotypes and files containing sequences for further analysis. To evaluate reliability of batch processing with the shell script, the similarity of sequences generated by the script and the sequences processed manually was analyzed by MEGA 6.0 software. Results The shell script accomplished all functions. The analysis results by the shell script were identical to those by manual analysis. Conclusions Applying shell script saved time and resources in sequencing results analysis. Researcher will be able to concentrate more on further analysis.