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README.md

POEM is a pipeline which can predict operons and core operons from metagenomic genome/assembly or short reads. It can be run on most * NIX systems.

Requirements

This pipeline is available on Linux systems. Make sure that you have the following installed

  1. Anaconda for Python 2.7

  2. conda

    make sure to add path of conda to $PATH environment variable

Installation

$ git clone https://github.com/Rinoahu/POEM

$ cd ./POEM

$ bash ./install.sh

The installation script calls conda to install all the necessary python packages and software. PS: POEM calls either Prokka or MetaGeneMark for gene predictions. If the users want to use MetaGeneMark, they must install it by themselves, because MetaGeneMark requires academic users to agree to an license before downloading. After installtation, make sure to add path of binary executable file gmhmmp in MetaGeneMark to $PATH environment variable.

Example

example directory contain a genome fasta file of E.coli, run runme.sh to test the pipeline

$ cd ./example

$ bash ./runme.sh eco.fasta

Usage

For short reads:

$ bash ./bin/run_poem.sh -f reads.fsa -a y

reads.fsa is single fasta file. If the reads are paired-end files in fastq or fasta format, 
use the fq2fa of IDBA_UD to convert them to  a single fasta file.

For genome/assembly:

$ bash ./bin/run_poem.sh -f genome.fsa -a n

genome.fsa is the genome/assembly fasta file.

Output

POEM will create a directory named read.fasta_output to save the results. The results include serveral file:

1.  input.fsa:
    The contigs or saffolds of IDBA-UD output in fasta format

2.  input.fsa_gmk_aa.fsa and input.fsa.gff:
    The fasta file of protein sequence and gff file generated by prediction of MetaGeneMark on input.fsa from step 1

3.  input.fsa.cdhit and input.fsa.cdhit.clstr:
    The cd-hit clustering results of input.fsa_gmk_aa.fsa from step 2

4.  input.fsa_aa.fsa:
    Filtered input.fsa_gmk_aa.fasta according to the clustering result of step 3

5.  input.fsa.flt.blast8:
    Blastp results in -m 8 tabular format of input.fsa_aa.fsa from step 4 against COG database

6.  input.fsa.flt.blast8.flt:
    Filtered blastp output of step 5. The hits with identity < 80% are removed

7.  input.fsa.cog:
    COG annotations for the protein sequences of step 4. The first row of this tab-delimited file is the gene number. The rest of the file contains 3 columns: col 1 is the gene identifier, chromosome|scaffold name and cluster id split by "$$", col 2 is the COG identifier and col 3 is the functional category of the genes, for example:

        uniq gene 3679
        gene	cog	annot
        CPIBHFJP_04443|Prodigal:2.6|3106475_aa|-|4659209|4659544$$gi|985000614|gb|CP014225.1|$$>Cluster_3671$$*	COG1455	:G::Phosphotransferase system cellobiose-specific component IIC::Carbohydrate transport and metabolism
        CPIBHFJP_04442|Prodigal:2.6|3106198_aa|-|4658880|4659158$$gi|985000614|gb|CP014225.1|$$>Cluster_3846$$*	COG1455	G::Phosphotransferase system cellobiose-specific component IIC::Carbohydrate transport and metabolism
        CPIBHFJP_04441|Prodigal:2.6|3105940_aa|-|4658384|4658734$$gi|985000614|gb|CP014225.1|$$Orphan$$Orphan	COG1447	G::Phosphotransferase system cellobiose-specific component IIA::Carbohydrate transport and metabolism
        ...


8.  input.fsa.locus:
    This file records the gene's locus on genome. This file contains 5 columns: col 1 is gene identifier and chromosome|scaffold name split by "$$", col2 is the chromosome|scaffold name, col3 is the strand of the gene, col 4 and 5 are the start and end of the gene. For example:

	CPIBHFJP_00001|Prodigal:2.6|772_aa|+|255|857$$gi|985000614|gb|CP014225.1|       gi|985000614|gb|CP014225.1|     +       255     857
	CPIBHFJP_00002|Prodigal:2.6|1014_aa|+|883|1308$$gi|985000614|gb|CP014225.1|     gi|985000614|gb|CP014225.1|     +       883     1308
	CPIBHFJP_00003|Prodigal:2.6|1165_aa|+|1586|1693$$gi|985000614|gb|CP014225.1|    gi|985000614|gb|CP014225.1|     +       1586    1693
	...


9.  input.fsa.adjacency and input.fsa.operon:
	This file is the predicted operonic adjacent genes. This file contains 11 columns: col 1 is the gene 1's id; col 2 is the chromosome|scaffold id where gene 1 is located; col 3-5 are strand, start and end of gene 1; col 6 is gene 2's id; col 7 is the chromsome|scaffold id where gene 2 is located; col 8-11 are strand, start and end of gene 2. For example:

		CPIBHFJP_00001|Prodigal:2.6|772_aa|+|255|857$$gi|985000614|gb|CP014225.1|       gi|985000614|gb|CP014225.1|     +       255     857     CPIBHFJP_00002|Prodigal:2.6|1014_aa|+|883|1308$$gi|985000614|gb|CP014225.1|     gi|985000614|gb|CP014225.1|     +       883     1308    False
		CPIBHFJP_00002|Prodigal:2.6|1014_aa|+|883|1308$$gi|985000614|gb|CP014225.1|     gi|985000614|gb|CP014225.1|     +       883     1308    CPIBHFJP_00003|Prodigal:2.6|1165_aa|+|1586|1693$$gi|985000614|gb|CP014225.1|    gi|985000614|gb|CP014225.1|     +       1586    1693    False
		...


10.  input.fsa.operon:
	The predicted full operons. This file contains 3 columns: col 1 is the ids of operonic genes split by --> or <--, arrow stands for the strand of gene; col 2 is the COG ids of the operonic genes; col 3 is the COG annotation of the operonics genes split by --> or <--. For example:
		
		gene_id COG_id  annotation
		CPIBHFJP_00007|Prodigal:2.6|2673_aa|+|3821|3946$$gi|985000614|gb|CP014225.1|-->CPIBHFJP_00008|Prodigal:2.6|2862_aa|+|3961|4182$$gi|985000614|gb|CP014225.1|-->CPIBHFJP_00009|Prodigal:2.6|3078_aa|+|4218|4484$$gi|985000614|gb|CP014225.1|-->CPIBHFJP_00010|Prodigal:2.6|3373_aa|+|4484|4867$$gi|985000614|gb|CP014225.1|-->CPIBHFJP_00011|Prodigal:2.6|3449_aa|+|4887|5078$$gi|985000614|gb|CP014225.1|        unknown-->unknown-->unknown-->COG3001-->COG3001 unknown::unknown-->unknown::unknown-->unknown::unknown-->G::Fructosamine-3-kinase::Carbohydrate transport and metabolism-->G::Fructosamine-3-kinase::Carbohydrate transport and metabolism
		CPIBHFJP_00036|Prodigal:2.6|20592_aa|+|29823|30533$$gi|985000614|gb|CP014225.1|-->CPIBHFJP_00037|Prodigal:2.6|21036_aa|+|30538|31215$$gi|985000614|gb|CP014225.1|-->CPIBHFJP_00038|Prodigal:2.6|21527_aa|+|31230|31937$$gi|985000614|gb|CP014225.1|-->CPIBHFJP_00039|Prodigal:2.6|21840_aa|+|31937|32485$$gi|985000614|gb|CP014225.1|-->CPIBHFJP_00040|Prodigal:2.6|22830_aa|+|32495|33661$$gi|985000614|gb|CP014225.1|-->CPIBHFJP_00041|Prodigal:2.6|23958_aa|+|33634|35169$$gi|985000614|gb|CP014225.1|-->CPIBHFJP_00042|Prodigal:2.6|24099_aa|+|35169|35822$$gi|985000614|gb|CP014225.1|-->CPIBHFJP_00043|Prodigal:2.6|25233_aa|+|35889|37196$$gi|985000614|gb|CP014225.1|   COG0398-->unknown-->COG0398-->COG2128-->COG4134-->COG4135-->COG4136-->COG2897   S::Uncharacterized membrane protein YdjX, TVP38/TMEM64 family, SNARE-associated domain::Function unknown-->unknown::unknown-->S::Uncharacterized membrane protein YdjX, TVP38/TMEM64 family, SNARE-associated domain::Function unknown-->P::Alkylhydroperoxidase family enzyme, contains CxxC motif::Inorganic ion transport and metabolism-->R::ABC-type uncharacterized transport system YnjBCD, periplasmic component::General function prediction only-->R::ABC-type uncharacterized transport system YnjBCD, permease component::General function prediction only-->R::ABC-type uncharacterized transport system YnjBCD, ATPase component::General function prediction only-->P::3-mercaptopyruvate sulfurtransferase SseA, contains two rhodanese domains::Inorganic ion transport and metabolism


11. input.fsa.cog_adjacency:
    This file contain two parts: The first part converts operonic adjacency to COG id adjacency and statistic of the COG id adjacency. The second part evaluate the core operon and its closest real operon.

		##############################
		# COG adjacency
		##############################
		geneA geneB cA cB cAB cA+cB-cAB cAB/(cA+cB-cAB)
		COG3188 COG3539 11 32 7 36 0.194444444444
		COG2801 COG2963 16 16 11 21 0.52380952381
		COG1662 COG3677 10 9 8 11 0.727272727273
		...

		##############################
		# evalute the core operon
		##############################
		predict_operon  real_operon_name        real_operon_cog Precise Recall  F1      predict_annot   real_annot
		COG0601$$COG1173        sso:SSO3058$$sso:SSO3059        COG0601$$COG1173        1.000000        1.000000        1.000000        ABC-type dipeptide/oligopeptide/nickel transport system, permease component$$ABC-type dipeptide/oligopeptide/nickel transport system, permease component        ABC-type dipeptide/oligopeptide/nickel transport system, permease component$$ABC-type dipeptide/oligopeptide/nickel transport system, permease component
	COG1129$$COG1172        eco:b1513$$eco:b1514$$eco:b1515$$eco:b1516$$eco:b1517$$eco:b1518$$eco:b1519     COG1129$$COG1172$$COG1172$$COG1879$$COG1830$$COG1359$$COG4106   1.000000        0.428571        0.600000        ABC-type sugar transport system, ATPase component$$Ribose/xylose/arabinose/galactoside ABC-type transport system, permease component    ABC-type sugar transport system, ATPase component$$Ribose/xylose/arabinose/galactoside ABC-type transport system, permease component$$Ribose/xylose/arabinose/galactoside ABC-type transport system, permease component$$ABC-type sugar transport system, periplasmic component, contains N-terminal xre family HTH domain$$Fructose-bisphosphate aldolase class Ia, DhnA family$$Quinol monooxygenase YgiN$$Trans-aconitate methyltransferase
		...




12. input.fsa.core_cog_adjacency:
    The core COG adjacency is the same as the first part of file from step 11. This file is tab delimited.

		geneA   geneB   cA      cB      cAB     cA+cB-cAB       cAB/(cA+cB-cAB)
		COG1173 COG0601 5       5       5       5       1.0
		COG1172 COG1129 8       8       6       10      0.6
		COG2963 COG2801 16      16      11      21      0.52380952381
		COG1662 COG3677 10      9       8       11      0.727272727273


13. input.fsa.core_network.sif, input.fsa.core_node.tab:
    Network, node attribute and edge attribute extracted from step 11 for cytoscape visualization. Figure 1 shows an example to view the core operons by cytoscape.

Visualization of core operons Figure 1: visualization of core operons in cytoscape

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