I have human WGS raw reads and paired-end data. Illumina Hi-Seq 2000 was used for sequencing.
I want to assemble this genome either de-novo or reference-based. I want to assemble it chromosome-specifically so I can only focus on my chromosome of interest. That will save a lot of time for me. And my other goal is to extract a specific gene sequence for it.
So,
1. Please recommend any standard pipelines or literature to me.
2. Can I extract a specific gene or protein sequence from it? If yes, please suggest me the methods for that too.
A big thank you in advance. :)
Hi Pratanu Kayet
First of all, your assembly will be done on a personal workstation (pc or laptop) or a supercomputing cluster or a cloud server? Note that to assemble a human chromosome, for example chromosome 14 (107,043,718 bp) the GRASShopPER assembler requires 82 GB RAM. Here is the download site: https://bio.cs.put.poznan.pl/programs/5a6e3ec4433e908bbb000001
Reference: https://doi.org/10.1371%2Fjournal.pone.0202355
Otherwise, you could try MEGAHIT (https://github.com/voutcn/megahit) which is a memory efficient assembler.
Regarding your second question, you can extract a specifiq sequence by parsing and indexing with SAMtools, is pretty easy if you have the target sequence ID or header.
If you want to extract an unnanotated sequence, I recommend you an a strategy that I've been performed several times that consists of scanning the coding sequences of the (meta)genome (assembled sequences followed by ORF calling) with the specific HMM profile of the protein of interest and in a few minutes you can find candidate sequences with nice match odds. Feel free to email me [email protected] if you want more information
Good luck!
José Germán Serrano-Gamboa
@ Assembly will be done in a Linux workstation.
- Badges
- Science topic