Sequence Analysis Tools

Basic Local Alignment search tool BLAST

(http://blast.ncbi.nlm.nih.gov/Blast.cgi).

This tool was established to discover (local) homology between two

sequences. Protein and nucleotide sequence databases can be used for a

given sequence of interest. This program calculates the statistical

significance of an alignment (Altschul, Gish et al. 1990).

The BLAST algorithm has many variation; BLASTN, BLASTP, BLASTX,

TBLASTN, mega BLAST and psi-BLAST. These different algorithms use

are according to the query input (nucleotide, protein or translated sequences)

with searches against a vast number of organism sequences.

MUMmer (http://www.tigr.org/software/mummer)

MUMmer 3.0 is open-source software that enables genome sequence

comparison of large genomes. MUMmer can align incomplete genomes

with the system. The graphical viewing tools afford different ways to

analyse genome alignments (Kurtz, Phillippy et al. 2004).

Artemis (http://www.sanger.ac.uk/Software/Artemis/v8/)

Artemis is a DNA sequence viewer and annotation tool that allows

visualisation of sequence features and the results of analyses within the

context of next generation data.

CLUSTALW (http://www.ebi.ac.uk/Tools/msa/clustalw2/)

CLUSTALW (1.83) is one of the most powerful programs used to achieve

multiple sequence alignments. This program allows the presentation of

multiple nucleotide and protein sequence alignments (Larkin, Blackshields

et al. 2007).

MUSCLE (http://www.drive5.com/muscle/)

MUSCLE (v3.6) is a computer program most widely used in biology to

create multiple sequence alignments of proteins. MUSCLE uses different

algorithms including fast distance estimation and progressive alignment.

The accuracy and speed of the program is better than CLUSTALW, since

hundreds of sequences can be aligned in seconds (Edgar 2004).

FigTree (http://tree.bio.ed.ac.uk/software/figtree/)

FigTree (v1.3.1) is a program for graphical viewing of phylogenetic trees.

The program was designed to show summarized and annotated trees formed

FastTree (http://www.microbesonline.org/fasttree/)

FastTree (v2.1.7) is an open-source software construct, which can infer

maximum likelihood phylogenetic trees from alignments of nucleotide or

protein sequences. Millions of alignments can be done in a reasonable

amount of time and memory (Price, Dehal et al. 2010).

Geneious (http://www.geneious.com/)

By using Geneious (v7.1.3) software, one can analyse integrated protein and

DNA sequences, perform BLAST and get access to public databases. The

most powerful analysis that can be done using this software is the sequence

alignments manageability for both pair-wise and multiple sequence

alignments and visualization of the sequence alignments. The alignment

results can be viewed as phylogenetic trees.

OrthoMCL (http://www.orthomcl.org/orthomcl/?rm=orthomcl)

OrthoMCL (v1.4) is one of the most commonly used programs to perform

identification of orthologous groups. In addition, access to these groups is

extremely important for study gene/protein evolution and comparative

genomics and genome annotation.

All against All BLASTP between species and within species with Markov

Cluster algorithm methods can be performed to find all orthologous groups

with any recent paralogs. Ortholog analysis by using OrthoMCL can be

applied with two genomes or it can be extensive to cluster orthologs from

multiple species in order to constructing orthologous groups (Li, Stoeckert

Mauve (http://gel.ahabs.wisc.edu/mauve)

Mauve (v2.3.1) software is a powerful package applied to determine the

presence of rearrangements and horizontal transfer in a genome. It is used

for the identification and alignment of conserved genomic DNA (Darling,

Mau et al. 2004). Mauve alignments were used in this study to draw

comparison between whole genomes as well as examine the reasons of

rearrangements within genomes of E. faecium.

BRIG (http://sourceforge.net/projects/brig/)

The BLAST Ring Image Generator BRIG (v1.0) is a desktop application

written in Java 1.6. This application was used in genome comparisons and

generates a circular image for the genome. The comparison in this

application depends on the Basic Local Alignment Search Tool (BLAST)

and CGView for image rendering. For generating genomes maps in BRIG in

this study DNA or protein files were used.

MeV (http://www.tm4.org/mev.html)

Multi experiment Viewer MeV (v10.2) is a beneficial microarray data

analysis tool, including high-level algorithms for statistical analysis,

classification, clustering, visualization, and biological argument discovery

(Chu, Gottardo et al. 2008). MeV was used in this study for clustering

Unipro UGENE (http://ugene.unipro.ru)

UGENE (v1.11.5) is open-source software that can be used as a

multiplatform software. It offers visualization of annotated genome

sequences, multiple sequence alignments and phylogenetic trees

(Okonechnikov, Golosova et al. 2012). In this study UGENE software was

used to identify and map the repetitive units in the genomes.

Phenolink (http://bamics2.cmbi.ru.nl/websoftware/phenolink/)

Phenolink is a web-tool to identify genetic links between phenotypes. It uses

~omics technologies that connect phenotypes with high-throughput

molecular biology information. The purpose is to see through cellular

mechanisms underlying an organism's phenotype (Bayjanov, Molenaar et al.

2012). A default parameter was used to identified E. faecium phenotypes.

CRISPRs Finder (http://crispr.u-psud.fr/Server/CRISPRfinder.php)

CRISPRFinder is a free access web service. CRISPRs stands for Clustered

regularly interspaced short palindromic repeats. Five tools are available in

CRISPRs Finder, which can be used for:

1. Detecting very short CRISPRs that consist of one or two motifs.

2. Identifying highly conserved regions (DR) and extracting similarly sized

unique sequences, which lie between the DRs called spacers.

3. Obtaining the AT-rich leader sequence, which flanks the CRISPR cluster

on one side.

5. To identify the highly conserved regions (DR) are present in other

prokaryotic sequenced genomes (Grissa, Vergnaud et al. 2007).

Island Viewer (http://www.pathogenomics.sfu.ca/islandviewer)

IslandViewer is a freely accessible web service that provides detection of

gene clusters likely to be of horizontal origin, called Genomic islands (GIs).

These clusters contain genes such as virulence, antibiotic resistance or other

important adaptation genes. IslandViewer uses a graphical interface that

allows easy viewing and the island data of both the chromosome and the

gene level can be downloaded. The server uses three methods to identify the

GI regions. IslandPick; comparative genomic GI prediction method to

advance stringent data sets of GIs and non-GIs, SIGI-HMM; This method

measures codon usage to identify possible GIs by using Hidden Markov

Model (HMM). Finally, IslandPath-DIMOB; this method visualises several

common characteristics of GIs such as abnormal sequence composition or

the occurrence of genes that are functionally related to mobile elements

(Langille and Brinkman 2009).

PHAST (http://phast.wishartlab.com)

PHAST is a fast web server used to distinguish, annotate and graphically

present prophage sequences and prophage features within bacterial genomes

IS Finder (https://www-is.biotoul.fr// )

IS Finder is a database provides a list of insertion sequences elements

isolated from Eubacteria and Archaea. The IS elements in this database are

defined in individual files which contains their general features such as

name, size and family plus their DNA and protein sequences. In addition,

for the comparison an on-line BLAST search is available.