Thanks Sean. You are right. That's why i said i will use RNAseq PE reads as SE reads because relative position in RNAseq will be different from those in DNAseq. But still we also need to consider intron/extron structure issues uniq to RNAseq as mentioned by akoik063. Cheers
Can Rnaseq Reads Be Used For Genome Assembly
Hi There,
I have been doing genome assembly with one pair-end library (insert length = 500) for a non-model plant species using Velvet. The assembly is very fragmented.
Since i also have pair-end RNAseq sequencing data, I am wondering if i can use RNAseq data to improve genome assembly.
My questions are: (1) Does it make sense to map transcriptome assembly to genome assembly and join genome scaffolds/contigs spanned by the same transcripts?
(2) can I try using RNAseq reads data for the same purpose? I mean using RNAseq reads as genomic sequence reads to do de novo assembly. Considering RNAseq PE reads are from alternatively spliced transcripts, I will use them as single-end reads when doing genome assembly.
I will appreciate it very much if someone can point out whether these ideas are reasonable or not or give me additional suggestions.
Kind Regards,
Lhl
5 answers
Came across some more tools (some included from above)
AGOUTI: improving genome assembly and annotation using transcriptome data https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4952227/
Rascaf: Improving Genome Assembly with RNA Sequencing Data https://www.ncbi.nlm.nih.gov/pubmed/27902792
PEP_scaffolder: using (homologous) proteins to scaffold genomes https://academic.oup.com/bioinformatics/article/32/20/3193/2196523
SCUBAT (Scaffolding Contigs Using BLAT And Transcripts) https://github.com/elswob/SCUBAT
L_RNA_scaffolder: scaffolding genomes with transcripts https://bmcgenomics.biomedcentral.com/articles/10.1186/1471-2164-14-604
This is NOT an evidence-based answer and represents only intuition, so I hope someone else has more insight. You propose an interesting idea, but I suspect that you are better off doing further genomic sequencing (with potentially different library prep or even technology). The RNA-seq single-end reads may, themselves, be spliced, making including them problematic. Including the RNA-seq as paired-end is also difficult since the insert size distribution is not well-understood given splicing.
• 135 views
•
link
written
by
Sean Davis
These two tools are supposed to perform this:
L_RNA_Scaffolder http://www.biomedcentral.com/1471-2164/14/604
SCUBAT https://github.com/elswob/SCUBAT
edit* I misread, you want to use RNA-seq reads. These tools used assembled transcripts to attempt to scaffold.
• 0 views
•
link
written
by
Damian Kao
Hi,
I too have the same question.Actually I am looking for a denovo assembly tool that can assemble a meta-transcriptome data(paired-end sequence(insert size=300) **consists of mixed sequence reads of multiple species in a microbial community.
And in search of that I came across a METAVELVET,a de novo metagenome assembly and I am not sure whether this works well with my data?
Any suggestions please.
• 0 views
•
link
written
by
bambus0725 •
I would be hesitant to use RNA-Seq reads for this purpose - genome assembly - because you cannot be certain that the reads are contiguous with respect to a complete genome. I would be much less hesitant to throw into the assembly process RNA-Seq reads from genes that are expressed as a single exon.
You noted that you're working with a non-model plant genome, but can you align reads to a completed plant genome? Not all available plant genomes are for model species. This may allow you to order reads more efficiently and with greater confidence than with the RNA-Seq reads. Or, it may give you one assembly of the genome that you can use. The RNA-Seq assembly can be another. My point is I found synteny very powerful when scaling from Arabidopsis to soybean.
• 205 views
•
link
written
by
Larry_Parnell
Hi, I found the following tools: * http://bmcresnotes.biomedcentral.com/articles/10.1186/s13104-016-1903-z#Sec2 * http://www.fishbrowser.org/software/PEP_scaffolder/
Or does any one know a better tools?
Mic
Log in to answer this question.
More posts like this
-
Calculating alignment/mapping time
written by kumars.sv •
-
Paired-end and single-end reads combined during RNAseq - acceptable?
written by iktu23 •
-
Should I do further scaffolding or gap-filling after hybrid assembly with DBG2OLC?
written by pgaiero •
-
Why mate-Paired end, Paired-end and Single End reads library to be combined for assembling?
written by saranpons3
-
Large Plant genome Assembly using ABYSS
written by mehtaaditya99 •
-
A Question About Hybrid/Mix -Genome Assembly
written by Lhl
-
Does It Make Sense To Merge Oases Multiple-Kmer Assemblies
written by Lhl
-
Is It Reasonable To Combine Both Genome And Transcriptome Sequences To Get A Larger Assembly ??
written by Lhl
-
How To Distinguish Between Heterozygous Or Duplicated Allels
written by Leszek
-
Genome Assembly With Physical Map
written by Plantae
Highly fragmented assemblies with velvet are not uncommon. Before I can advise furhter: 1) Is this illumina sequencing? Is it MiSeq by any chance? What is the coverage? 2) Have you tried Velvet Optimiser?
And to answer your question directly, I wouldn't use RNASeq reads for genome assembly. Think about it, if it's a eukaryote, it could have introns. Even if it's not, we are talking about gene duplications and repetitive regions...
Thanks akoik063.
It is Illumina-highseq sequencing. About the coverage, Velvet produced the following message (k=57) 'Median coverage depth = 4.293333 Final graph has 5301633 nodes and n50 of 1063, max 70820, total 613220706, using 78039534/136079432 reads'. I mapped reads to the assembly and did some calculations and got average coverage == 45. I have NOT tried Optimiser, i simply tried multiple k-mers and found k=57 gave largest N50.