Background Place bZIP proteins characteristically harbor a highly conserved bZIP website with two structural features: a DNA-binding fundamental region and a leucine (Leu) zipper dimerization region. With the availability of these legume genome sequences the users of the bZIP transcription element family were systematically investigated GSK690693 and analyzed with this study. We recognized all legume bZIP genes and analyzed their bZIP website GSK690693 sequences gene structure and GSK690693 additional MEME motifs which was in agreement with and supported the phylogenetic classification. Then we expected the DNA-binding-site specificity and dimerization properties of the legume bZIP proteins. We also investigated the effect of the two legume-lineage WGDs and tandem duplication within the expansion of the legume bZIP gene family. By analyzing their expression profiles legume GSK690693 bZIP genes constitutively or specifically expressed in different cells and seed developmental phases were identified as well as candidate legume bZIPs responsive to drought and salt stresses. Methods Recognition of bZIP transcription factors in six legume genomes All genomic sequences and annotated proteins of the six legumes were downloaded from ftp://ftp.jgi-psf.org/pub/compgen/phytozome/v9.0/Gmax/ (and the six legume genomes were aligned using ClustalX 2.0  with space opening and space extension penalties of 10 and 0.1 respectively. The phylogenetic tree was reconstructed by the maximum likelihood (ML) method using the PhyML 3.0 software . JTT?+?G was selected while the best model for constructing the phylogenetic tree from the Akaike info criterion implemented in ProtTest 3.0 . Bootstrap ideals from 100 replicates were indicated at each node. MEGA5  was used to show the tree. Structure of bZIP genes The positional info of both the gene sequence and the related coding sequence were loaded into the gene structure display server v2.0 (http://gsds.cbi.pku.edu.cn/)  to obtain info within the intron/exon structure. The coordinates of the bZIP website in each protein were recalculated into the coordinates in gene sequence and presented in gene structure. We used Genewise  to analyze the intron distribution pattern and intron splicing phase FAS within the basic and hinge regions of the bZIP domains in the six legumes. Detection of additional conserved motifs To identify additional conserved motifs outside the bZIP website of legume bZIP transcription factors we used the Multiple Em (Expectation Maximization) for the Motif Elicitation tool (MEME version 4.9.1 http://meme.nbcr.net/meme/) . The limits for maximum width minimum width and maximum quantity of motifs were specified as 50 10 and 100 respectively. Fifty motifs were finally confirmed because of their low e-values (<1e-200). The motifs were numbered according to the order displayed in MEME and were considered as group-specific signatures for his or her presence of high rate of recurrence in the given groups. Detection of duplicated genes and estimation of synonymous (Ks) and nonsynonymous (Ka) substitutions per site and their percentage The duplicated gene pairs derived from segmental duplication were recognized in the legume genomes based on the method from your Flower Genome Duplication Database . An all-against-all BLASTP assessment (e-value: 1e-5) offered the gene pairs for syntenic clustering determined by MCScan (using default settings: MATCH_SCORE: 50 MATCH_SIZE: 5 Space_SCORE:-3 E_VALUE: 1E-05) (http://chibba.agtec.uga.edu/duplication/mcscan). Tandem duplication arrays were recognized using BLASTP having a threshold of e?10?20 and one unrelated gene among cluster users was tolerated while described in . Pairs from segmental or tandem duplications were used to estimate Ka Ks and their percentage. Amino acid sequences from segmentally or tandemly duplicated pairs had been first aligned and guided and moved right into a cDNA sequences alignment using in-house Perl scripts. Then your software program KaKs_Calculator was utilized to compute Ka and Ks beliefs for each set following YN model . Appearance evaluation of legume bZIP genes For different tissue/body organ and seed developmental levels the normalized matters for bZIP genes from RNA-seq had been.