howto(obsolete)‎ > ‎

HOSOMICHI HLA ANALYSIS TOOLS

Introduction

In this section, we would like to introduce p-Galaxy workflows to perform ported "HLA ANALYSIS TOOLS" developed by Dr. Hosomichi of Division of Human Genetics, National Institute of Genetics of Japan.
Human Leukocyte Antigen (HLA) is a group of genes that are extremely polymorphic among individuals and populations. The HLA has been associated with more than 100 different diseases, mostly autoimmune diseases. The workflows are capable to perform the HLA typing for medical research, and population genetics.

References:

Hosomichi et al. Phase-defined complete sequencing of the HLA genes by next-generation sequencing. BMC Genomics 2013, 14:355.

Hosomichi et al. A simple and rapid 96-well plate-based complete sequencing method of the HLA-B (Submitted).

Nagasaki et al. DDBJ Read Annotation Pipeline: A Cloud Computing-Based Pipeline for High-Throughput Analysis of Next-Generation Sequencing Data. DNA Res. 2013;20(4):383-90.


Preparing User Account in Common with DDBJ Pipeline

Originally, the Galaxy requires your mail address and password. P-Galaxy also requires mail address, which is submitted with user account of DDBJ Pipeline. Therefore if you do not have the pipeline account, you should have the account to use P-Galaxy. To have the account, please read described part in the manual.

When you have user account of the DDBJ Pipeline, start up P-Galaxy from P-Galaxy website OR from the link ("step-2/Workflow/Genome...") in the left side menu of basic analysis part of DDBJ Pipeline.




When if you can see P-Galaxy top page...

 

You should check you are already login or not from "top menu/User". 
If you are not login, set e-mail address and password (submitted with the pipeline account).


Login window is appeared. 


Data Transfer

In this section, how to transfer FASTQ files to HLA typing is described.
Mapping of genotype is performed using BWA software, thus FASTQ files should be included reads by illumina.

The step of data transfer to "galaxy" directory in the ftp server (pdata.nig.ac.jp) is mostly same as the illustration of section "5-2-2 Upload your files via FTP" using FTP client(FileZilla, Cyberduck, etc).



Look to p-galaxy.ddbj.nig.ac.jp, you can find "Get Data/Upload File from your computer" in left side tools of P-Galaxy window.
If the data has been uploaded, you can see the data name will be appeared in center frame of P-Galaxy.
Press "Execute" button to transfer the data to P-Galaxy.


The loaded data will be appeared in "history" area in right side of P-galaxy window.



Setting Up Workflows

Choose the menu of "Published workflows" of "Shared Data" menu in c
entrally above of the window.



The "Published Workflows" flame is opened.

Choose "import" in the menu of 
"HLAwfSeparate1" and "HLAwfSeparate2".

The workflow "HLAwfSeparate1" performs from trimming reads to building HLA sequences exchanged polymorphisms between mapped reads and HLA genome.
The second, 
"HLAwfSeparate2" makes the typing lists from polymorphisms between HLA sequences HLAwfSeparate1 and HLA genome. 

   


Running Workflow "HLAwfSeparate1"

After import HLAworkflows, click "workflow/ワークフロー" in the center of the upside menu, imported workflows will be appear. Choose "Run" from "imported: HLAwfSeparate1" menu.



The procedure of the workflow is appeared in the Galaxy window, set the forward/reverse reads and choose HLA (A/B/C/DPB1/DQB1/DRB1) genome.
At the end of the procedure, a button of "Run workflow".


   


HLA sequences will be generated in History flame in the left side of P-Galaxy window.

Running Workflow "HLAwfSeparate2"

Following "HLAwfSeparate1", "HLAwfSeparate2" makes typing results from respective HLA sequence.
   

NOTICE ABOUT AN ERROR

Some FASTQ files make an error in the step of the finding haplotypes of HLAwfSeparate1 procedure. The cause of this error is that HLA sequences are split by few reads mapping to the reference HLA genome. The mapping methods is BWA MEM, which allows loose mapping, but some read set make that (please check matching to the reference). Recently we are planning to decrease the occurrence of this error.


Sample FASTQ Files for Trial HOSOMICHI HLA ANALYSIS TOOLS

K-0328_S66_L001_R1_001.fastq (forward) and K-0328_S66_L001_R2_001.fastq (reverse)


Choose HLA-B as reference genome with above FASTQ data.




Comments