Bacterial de novo genome assembly

Question

[Paul]

I have pair end Fastq files from illumina (Fast1.fastq and Fast2.fastq). Now, how should I proceed further with de-novo genome assembly? I'am completely new to this field of assembling genome sequences? Which open-end tools can be used?

Please help me with the tools and tutorials which can be used for de-novo assembly of sequences.

[Titus]

Hi , You can read this article to start : http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0017915

Best

[dago]

what did you try? There are plenty of tutorials out there. What organism do you have? Bacteria?

[Paul]

Yes it is for Bacteria

[Matteo Schiavinato]

I think we need a little more details to help you. If you want to learn general procedures, well the literature is there for you as it was for us!

[Paul]

Ya general procedures with the softwares available

[Titus]

I spoke with friend who work on meta genomic Bacteria data and to him the most use software for bacteria assembly is SPADes.

Best

Answer 1

[Brian Bushnell]

If you download the BBMap package, there is a (hopefully) helpful guide in bbmap/docs/guides/PreprocessingGuide.txt

For isolate bacterial assembly with SPAdes, I recommend:

1) Adapter-trimming (you can do quality-trimming at the same time; I suggest a low cutoff, such as Q10)

2) Artificial contaminant filtering

3) Human contaminant removal

4) Error correction

5) Paired-read merging

...then assemble with SPAdes, using both the merged and unmerged reads.

[dago]

check assembly quality with QUAST

[Paul]

Thanks a lot for the detailed explanation. Is there any best tool to do the bacterial genome assembly in windows?

[WouterDeCoster]

You will have so much more options if you use Linux... Windows is a great operating system, but not for bioinformatics.

[dago]

Look these: Best software to assemble bacterial genomes And also a list of tools available is here. You can check the one that work in windows amongst them.

[Joe]

I don't know of many off-hand. CLC Genomics Workbench will do assembly, but it isn't free. It's a decent one-stop-shop for pretty much everything however (and is GUI based if you're commandline averse).

Answer 2

[Joe]

The general procedure I follow is:

1. Quality control your data before you do anything else. FastQC is a go-to tool for this.
2. Trim sequences to remove adapters etc if necessary. (See Seqtk, sickle, cutadapt, trimgalore and other such tools)
   • It may make sense to filter the reads out at this stage and throw away anything that is garbage.
3. Assemble (See SOAP, Velvet, SPAdes)
4. Re-map the reads to get coverage statistics etc (optional but advised). (See, Bowtie2, BWA, Qualimap etc)
5. Optionally, if you have published reference genomes, you may wish to reorder contigs (See progressiveMauve/Mauve)

[Antonio R. Franco]

Not to forget that you can use a reference assisted assembly if you consider that a nice reference genome can be used for this purpose

[Joe]

Indeed :) OP asked for de novo specifically, but if reference guided is desirable I believe programs like MIRA will do it.

[Antonio R. Franco]

Many other programs do it. In particular, Velvet and SOAP can use reference genomes to assist a de novo assembly.

In my hands. the using of a trusted reference genome for assisting a de novo assembly, gave better results than a de novo assembly that was processed by Mauve afterwards