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ORFs were identified by using the EMBOSS software (version 5.0.0; emboss.sourceforge.net).net
All bioinformatics analysis was carried out using the EMBOSS software package and our previously published database KISS [15], [29].
The genome sequences of M. tuberculosis strains H37Rv (GenBank: AL123456) [29] and CDC1551 (GenBank: AE000516) [30] were analysed using the EMBOSS software (http://www.emboss.sourceforge.net).net
Sequence analysis was performed using the Emboss software suite [ 37], Clustal W [ 38] and 3DCoffee [ 39].
The identification of CpG islands and the plotting of GC contents were performed using the EMBOSS software suite [ 55].
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These were then aligned using the EMBOSS 'water' local alignment software (Rice et al., 2000) to the D. discoideum, Drosophila melanogaster (AAB58975.1), and Takifugu rubripes (AAD15839.1) NF1 orthologues and the Saccharomyces cerevisiae Ira1p protein (NP_009698.1).
Intron/exon boundaries were identified by alignment of clover or legume orthologue cDNA with clover genomic sequence orthologues, or publicly available genomic legume databases such as M. truncatula or Lotus japonicus genomic sequence using the EMBOSS (European Molecular Biology Open Software Suite) est2genome programme.
Alignments of the parent genes and CG proteins were generated using the Emboss Needle program (http://emboss.sourceforge.net/) with default options.
GC content was calculated using the EMBOSS geecee program.
(G + C) content was calculated using the eMBOSS geecee program http://inn-temp.weizmann.ac.il/cgi-bin/emboss/geecee[ 63].
Sequences were translated to amino acids using the Emboss Transeq web application [ 87].
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