Question: Merge Paired-End Reads
9
gravatar for NicoBxl
4.6 years ago by
NicoBxl4.0k
Belgium
NicoBxl4.0k wrote:

Hi,

How can I merge two paired end fastq (R and L) to give a single fastq file ? For information, the sequencing run is 72 bp long and it contains a majority of small RNA (miRNA,...) so a lot of paired end reads will overlap.

For example here's two paired reads :

@HWUSI-EAS529:41:FC62YHFAAXX:8:1:7969:1330 1:N:0:GCCAAT
CTACGAAAGGGCACTTGGAATTCTCGGGTGCCAAGGAACTCCAGTCACGCCAATATCTCGTATGCCGTCTTCTGCT
+
IIIIIIIHIIHIIIIIIHHIIIHGIIIIEIIIIIIEIIHIIIIIIIIIIIHIIIIIBHIHIIHGIGIEGHHEGEEH


@HWUSI-EAS529:41:FC62YHFAAXX:8:1:7969:1330 2:N:0:GCCAAT
AGTGCCCTTTCGTAGGATCGTCGGACTGTAGAACTCTGAACGTGTAGATCTCGGTGGTCGCCGTATCATTAAAAAA
+
IIIIIIIIIIIIIIIIIIIIIIIDHIGIIIHIIIGHGIIIIIIIHHIHIIIIIIIIIHIIIIIIIIHIIGIIIIHI

I find the adapter in the first one :

Code:

EMBOSS_001         1 CTACGAAAGGGCACTTGGAATTCTCGGGTGCCAAGGAACTCCAGTCACGC     50
                                    |||||||||||||||||||||              
EMBOSS_001         1 ---------------TGGAATTCTCGGGTGCCAAGG--------------     21

EMBOSS_001        51 CAATATCTCGTATGCCGTCTTCTGCT     76

EMBOSS_001        22 --------------------------     21

but not in the second one ...

But I effectively found the overlap between the right read and the left read (using the reverse complement of it)

EMBOSS_001         1 --------------------------------------------------      0

EMBOSS_001         1 TTTTTTAATGATACGGCGACCACCGAGATCTACACGTTCAGAGTTCTACA     50

EMBOSS_001         1 -----------CTACGAAAGGGCACTTGGAATTCTCGGGTGCCAAGGAAC     39
                                |||||||||||||||                        
EMBOSS_001        51 GTCCGACGATCCTACGAAAGGGCACT------------------------     76

EMBOSS_001        40 TCCAGTCACGCCAATATCTCGTATGCCGTCTTCTGCT     76

EMBOSS_001        77 -------------------------------------     76

So my question is, how can I merge the two fastq files to produce a single fastq file ?

Thanks,

N.

paired merge rna fastq • 11k views
ADD COMMENTlink modified 4 months ago by midox90 • written 4.6 years ago by NicoBxl4.0k

Hi, I see that you had a similar case like me, so probably you can help me :) 

As I always do the miRNA analysis in single end I'm confused how to proceed when I have paired-end? Can you recommend me how to clean the reads and have them ready for analysis, particularly I cannot understand how and what is the relation of the reverse-compliment miRNA sequence in R2 read to the R1 set?
In summary my R1 read is containing 100nt - miRNA+barcode+smallRNAadapter+another adapter+polyA
my R2 is containing miRNA (reversed compliment to R1) + long adapter (or linker) + polyA

Thanks for any help in advance!

ADD REPLYlink written 10 months ago by manekineko60
2
gravatar for pmenzel
4.6 years ago by
pmenzel310
pmenzel310 wrote:

Maybe this program is suited for you: http://www.cbcb.umd.edu/software/flash/

ADD COMMENTlink written 4.6 years ago by pmenzel310

It's no available, the web page cannot be open. http://genomics.jhu.edu/software/FLASH/index.shtml

ADD REPLYlink written 3.5 years ago by litiancheng.gansu10

It's here now: http://ccb.jhu.edu/software/FLASH/

ADD REPLYlink written 2.9 years ago by matted5.9k
1
gravatar for Jeremy Leipzig
4.6 years ago by
Philadelphia, PA
Jeremy Leipzig16k wrote:

There is a decent program called stitch:[?] https://github.com/audy/stitch

I wrote a script called mergePairs that is very sensitive and incredibly slow:[?] http://code.google.com/p/standardized-velvet-assembly-report/source/browse/trunk/mergePairs.py

ADD COMMENTlink written 4.6 years ago by Jeremy Leipzig16k
1
gravatar for lelle
2.9 years ago by
lelle650
Berlin
lelle650 wrote:

As this was referenced from a duplicate question, I will add a newer tool to the list: PANDAseq

ADD COMMENTlink written 2.9 years ago by lelle650
1
gravatar for Andreas
14 months ago by
Andreas2.2k
Singapore
Andreas2.2k wrote:

One more: SeqPrep

Andreas

ADD COMMENTlink written 14 months ago by Andreas2.2k
0
gravatar for Stevelor
4.6 years ago by
Stevelor310
Stevelor310 wrote:

Either use Galaxy or use the single scripts

http://hg.notalon.org/galaxy/galaxy-central/src/7d9bb95caaa7/tools/fastq

HTH!

ADD COMMENTlink written 4.6 years ago by Stevelor310
1

uhhh which script?

ADD REPLYlink written 4.6 years ago by Jeremy Leipzig16k
0
gravatar for Gabriel R.
14 months ago by
Gabriel R.1.6k
Germany
Gabriel R.1.6k wrote:

If you wish to trim adapters and merge in a single step, you can the leeHom, we use it mainly to reconstruct ancient DNA sequences but it has broader uses as well:

http://nar.oxfordjournals.org/content/42/18/e141

It use a Bayesian maximum a posteriori approach that considers quality scores for both the adapter determination and the merging part. 

ADD COMMENTlink written 14 months ago by Gabriel R.1.6k
0
gravatar for midox
4 months ago by
midox90
Tunisia
midox90 wrote:

Hi,
So for an exact answer to this problem.
the R1.fq are the foward reads and the R2.fq are written in reverse-complement.
For example, if I want to create a single file from reads in R1.fq and R2.fq, I have to do "reverse-complement" of reads in R2.fq ??
am i right?
Thankyou

ADD COMMENTlink written 4 months ago by midox90

no response for this problem?

ADD REPLYlink written 4 months ago by midox90
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