De Novo Assembly Of A Bacterial Plasmid

Question

[mctomo]

Does anyone know what would be an easy and good way to assemble a bacterial plasmid genome (5-20kb) from Illumina reads that are 150 bases long? I have at least 1000x coverage. Some time ago I tried to use soap denovo and velvet but didn't get good results so I turned to mapping reads to a reference since I had the reference. Now I don't have a reference and would need de novo assembler. The plasmids don't contain repeats.

[Zev.Kronenberg]

Have you thought about trying a reference assisted assembler? Is this plasmid similar to other plasmids? I have really enjoyed using https://github.com/ibest/ARC?source=c ARC for assemblies.

[jigarnt]

Hi All,

I am trying to perform a de Novo assembly of the supposed plasmid that I have sequenced. In that case, which is the best tool/software which will give me the best results?

[Roye Rozov]

Hi jigarnt,

I recently posted about a new tool we developed called Recycler that is designed exactly for this purpose: Recycler for plasmid assembly

I hope it can help you. Cheers, R

[jigarnt]

**Getting this error: /Users/lindakohn/Desktop/tools/SPAdes-3.7.1-Darwin/bin/spades.py -k 21,33,55,77 --careful --only-assembler --pe<#>-12 <euro_plasmid_r1_paired.fastq euro_plasmid_r2_paired.fastq=""> --pe<#>-s1 <euro_plasmid_r1_unpaired.fastq> --pe<#>-s2 <euro_plasmid_r2_unpaired.fastq> -o Euro_plasmid_spades_output

-bash: syntax error near unexpected token `newline' what is wrong with the command?**

[mastal511]

Angle brackets < > indicate that some type of file name or value should be used in the command at that place, but they aren't part of the command itself.

If a Linux command spans several lines, you can use \ followed by return, to indicate that the command continues on the next line - that may be what is causing the error message.

Change your command to:

/Users/lindakohn/Desktop/tools/SPAdes-3.7.1-Darwin/bin/spades.py -k 21,33,55,77 \
--careful --only-assembler --pe1-1  euro_plasmid_r1_paired.fastq \
--pe1-2 euro_plasmid_r2_paired.fastq --pe1-s euro_plasmid_r1_unpaired.fastq \
--pe1-s euro_plasmid_r2_unpaired.fastq  -o Euro_plasmid_spades_output

Answer 1

[HG]

Hello, i used Spades for same types of study and got satisfactory result. You can have a look. http://bioinf.spbau.ru/spades

[mctomo]

I tried Spades and it worked beautifully! For a 7kb genome I got two contigs that overlap on both sides and make a circular genome. Thank you

Answer 2

[nicolasdierckxsens]

Hi, I developed a new de novo assembler for plastids and it should assemble the genome in one circular contig. I will upload the assembler in the next few weeks if you would be interested: https://github.com/ndierckx/NOVOPlasty

I could already upload a beta version next week, probably some bugs, but all tests were successful. I assembled 10 chloroplasts, all in one contig and within 30 min. The high coverage is no problem for this assembler and you don't need any reference. For the paper I assembled the chloroplast of Arabidopsis and rice, they were both 100 % accurate, so you should obtain a high quality assembly. I haven't tested it on plasmids but it's the same concept so should work..