How to align single end multiple lane data using Hisat2?
2
0
Entering edit mode
6 days ago
AANCHAL • 0

I have 4 lane , single end reads fastq files per sample that I need to align. How can I combine the 4 lanes to give as input?

I am trying to give the following command:

hisat2 -p 12 -x ./MM10_Index_HISAT/mm10 -U ./Rane_data/Acrolein-1_S4_L1_R1.fastq,./Rane_data/Acrolein-1_S4_L2_R1.fastq,./Rane_data/Acrolein-1_S4_L3_R1.fastq,./Rane_data/Acrolein-1_S4_L4_R1.fastq --rna-strandedness F -S ./HISAT_output

Also, how can I give all samples together in one command? I have 6 samples.

lanes Single reads • 437 views
ADD COMMENT
0
Entering edit mode

Please read the POSTING GUIDE before you Submit a new post. It's shown above the editor when you create a post. Also, please use the formatting bar (especially the code option) to present your post better. You can use backticks for inline code (`text` becomes text), or select a chunk of text and use the highlighted button to format it as a code block. If your code has long lines with a single command, break those lines into multiple lines with proper escape sequences so they're easier to read and still run when copy-pasted. I've done it for you this time.
code_formatting

ADD REPLY
0
Entering edit mode

Thank you. This was my 1st post. Will take care of it from next time.

ADD REPLY
0
Entering edit mode
6 days ago
ATpoint 47k

Yes, that is correct, see manual.

-U <r>

Comma-separated list of files containing unpaired reads to be aligned, e.g. lane1.fq,lane2.fq,lane3.fq,lane4.fq. Reads may be a mix of different lengths. If - is specified, hisat2 gets the reads from the “standard in” or “stdin” filehandle.

ADD COMMENT
0
Entering edit mode

I tried running my all sample files comma separated and it gave me an error. Is there a way to combine all samples?

ADD REPLY
0
Entering edit mode

and what is the error?

ADD REPLY
0
Entering edit mode
Error: Encountered internal HISAT2 exception (#1)

Command: /usr/local/hisat2/hisat2-2.1.0/hisat2-align-s --wrapper basic-0 -p 12 -x ./MM10_Index_HISAT/mm10 --rna-strandedness F -S ./HISAT_output -U ./Rane_data/Acrolein-1_S4_L1_R1.fastq,./Rane_data/Acrolein-1_S4_L2_ R1.fastq,./Rane_data/Acrolein-1_S4_L3_R1.fastq,./Rane_data/Acrolein-1_S4_L4_R1.fastq,./Rane_data/Acrolein-2_S5_L1_R1.fastq,./Rane_data/Acrolein-2_S5_L2_R1.fastq,./Rane_data/Acrolein-2_S5_L3_R1.fastq,./Rane_data/Acro lein-2_S5_L4_R1.fastq,./Rane_data/Acrolein-3_S6_L1_R1.fastq,./Rane_data/Acrolein-3_S6_L2_R1.fastq,.//Rane_data/Acrolein-3_S6_L3_R1.fastq,./Rane_data/Acrolein-3_S6_L4_R1.fastq,./Rane_data/Control-1_S1_L1_R1.fastq,./R ane_data/Control-1_S1_L2_R1.fastq,./Rane_data/Control-1_S1_L3_R1.fastq,./Rane_data/Control-1_S1_L4_R1.fastq,./Rane_data/Control-2_S2_L1_R1.fastq,./Rane_data/Control-2_S2_L2_R1.fastq,./Rane_data/Control-2_S2_L3_R1.fa stq,./Rane_data/Control-2_S2_L4_R1.fastq,./Rane_data/Control-3_S3_L1_R1.fastq,./Rane_data/Control-3_S3_L2_R1.fastq,./Rane_data/Control-3_S3_L3_R1.fastq,./Rane_data/Control-3_S3_L4_R1.fastq (ERR): hisat2-align exited with value 1

ADD REPLY
1
Entering edit mode

Acrolein-1_S4_L2_ R1.fastq,./Rane
-----------------^

There are spaces in filenames in your command. Are those copy-paste artifacts?

ADD REPLY
0
Entering edit mode

these are copy-paste artifacts

ADD REPLY
0
Entering edit mode

Did you put spaces after the commas? Also, are you completely sure that you want to align Acreolein-2 and Acreolein-3 together?

ADD REPLY
0
Entering edit mode

I want to do all in one go just like STAR if possible. STAR takes up all input files in the command line.

ADD REPLY
0
Entering edit mode
hisat2 -p 12 -x ./MM10_Index_HISAT/mm10 -U ./Rane_data/Acrolein-1_S4_L1_R1.fastq,./Rane_data/Acrolein-1_S4_L2_R1.fastq,./Rane_data/Acrolein-1_S4_L3_R1.fastq,./Rane_data/Acrolein-1_S4_L4_R1.fastq --rna-strandedness F -S ./HISAT_output

Right now this is my command line I am giving and it is also giving me error:

Error:

Encountered internal HISAT2 exception (#1)

Command:

/usr/local/hisat2/hisat2-2.1.0/hisat2-align-s --wrapper basic-0 -p 12 -x ./MM10_Index_HISAT/mm10 --rna-strandedness F -S ./HISAT_output -U ./Rane_data/Acrolein-1_S4_L1_R1.fastq,./Rane_data/Acrolein-1_S4_L2_
    R1.fastq,./Rane_data/Acrolein-1_S4_L3_R1.fastq,./Rane_data/Acrolein-1_S4_L4_R1.fastq 

(ERR): hisat2-align exited with value 1
ADD REPLY
0
Entering edit mode

I dont understand what I am doing wrong!

ADD REPLY
0
Entering edit mode

Can somebody help with the error?

ADD REPLY
0
Entering edit mode

If you're working on a cluster, check with your admin if you're running out of resources or something of that sort. Also, don't do this constant follow up commenting, it's rude.

ADD REPLY
0
Entering edit mode
6 days ago
swbarnes2 9.7k

With single end reads, you could also do something like

cat *.fastq.gz |  hisat2 -p 12 -x ./MM10_Index_HISAT/mm10 -U - --rna-strandedness F -S ./HISAT_output
ADD COMMENT
0
Entering edit mode

Also I downloaded the tar file of the mm10. The following are the files and contents in the mm10 folder.

total 3961472
-rw------- 1 a0malh01 unixuser  888467914 Mar 17  2016 genome.1.ht2
-rw------- 1 a0malh01 unixuser  663195880 Mar 17  2016 genome.2.ht2
-rw------- 1 a0malh01 unixuser       6119 Mar 17  2016 genome.3.ht2
-rw------- 1 a0malh01 unixuser  663195875 Mar 17  2016 genome.4.ht2
-rw------- 1 a0malh01 unixuser 1165549977 Mar 17  2016 genome.5.ht2
-rw------- 1 a0malh01 unixuser  675339278 Mar 17  2016 genome.6.ht2
-rw------- 1 a0malh01 unixuser          8 Mar 17  2016 genome.7.ht2
-rw------- 1 a0malh01 unixuser          8 Mar 17  2016 genome.8.ht2
-rwx------ 1 a0malh01 unixuser       1378 Mar 17  2016 make_grcm38.sh

Just wanted to make sure it is fine and not causing any error.

ADD REPLY
0
Entering edit mode

Is this file ok? It is not causing the error mentioned above?

ADD REPLY

Login before adding your answer.

Traffic: 2431 users visited in the last hour
Help About
FAQ
Access RSS
API
Stats

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.

Powered by the version 2.3.6