Singh Aditya, Bhatia Prateek
Advanced Paediatrics Centre, Post Graduate Institute of Medical Education and Research, Chandigarh, India.
J Biomol Tech. 2016 Dec;27(4):129-131. doi: 10.7171/jbt.16-2704-005. Epub 2016 Oct 17.
Sanger sequencing platforms, such as applied biosystems instruments, generate chromatogram files. Generally, for 1 region of a sequence, we use both forward and reverse primers to sequence that area, in that way, we have 2 sequences that need to be aligned and a consensus generated before mutation detection studies. This work is cumbersome and takes time, especially if the gene is large with many exons. Hence, we devised a rapid automated command system to filter, build, and align consensus sequences and also optionally extract exonic regions, translate them in all frames, and perform an amino acid alignment starting from raw sequence data within a very short time. In full capabilities of Automated Mutation Analysis Pipeline (ASAP), it is able to read "*.ab1" chromatogram files through command line interface, convert it to the FASTQ format, trim the low-quality regions, reverse-complement the reverse sequence, create a consensus sequence, extract the exonic regions using a reference exonic sequence, translate the sequence in all frames, and align the nucleic acid and amino acid sequences to reference nucleic acid and amino acid sequences, respectively. All files are created and can be used for further analysis. ASAP is available as Python 3.x executable at https://github.com/aditya-88/ASAP. The version described in this paper is 0.28.
诸如应用生物系统仪器之类的桑格测序平台会生成色谱图文件。通常,对于序列的1个区域,我们使用正向和反向引物对该区域进行测序,通过这种方式,我们有2条需要比对的序列,并在进行突变检测研究之前生成一个共有序列。这项工作既繁琐又耗时,特别是如果基因很大且有许多外显子的话。因此,我们设计了一种快速自动化命令系统,用于筛选、构建和比对共有序列,还可以选择提取外显子区域,在所有阅读框中进行翻译,并在极短的时间内从原始序列数据开始进行氨基酸比对。在自动突变分析管道(ASAP)的全部功能中,它能够通过命令行界面读取“*.ab1”色谱图文件,将其转换为FASTQ格式,修剪低质量区域,对反向序列进行反向互补,创建共有序列,使用参考外显子序列提取外显子区域,在所有阅读框中翻译序列,并分别将核酸和氨基酸序列与参考核酸和氨基酸序列进行比对。所有文件都会创建出来并可用于进一步分析。ASAP可作为Python 3.x可执行文件在https://github.com/aditya-88/ASAP上获取。本文所述版本为0.28。