Our results from SDS-PAGE analysis of abdominal muscles suggest the occurrence of muscle fiber rearrangement in both the premolt and postmolt stages [4]. The genes and molecular mechanisms of shrimp muscle growth have not received adequate scientific attentions to date. To gain a better understanding of shrimp growth and the underlying molecular mechanisms, we initiate a research project to identify muscle structural and regulatory genes by cDNA library and gene expression analysis. In a previous study, the abdominal muscle cDNA library of shrimp L.vannamei was successfully constructed [6]. Preliminary data analysis suggests that abundant and diverse transcripts are present in the cDNA library established by our laboratory. By degenerated PCR primer designing, we recently identified shrimp SUMO cDNA named LvSUMO-1 in L. vannamei. Expression of LvSUMO-1 mRNA suggest its role in the regulation of shrimp muscle protein degradation . In this research, we identified a novel gene encoding a putative DNA- binding protein, and further characterized its gene expression patterns in various shrimp muscle tissues and during the molt cycle. Results and Discussion LvDBP23 cDNA and Amino Acid Sequences analysis By using degenerated oligonucleotide DNA probes, we have identified muscle regulatory genes from L. vannamei muscle cDNA library. One of the interesting genes reported here is a novel DNA binding protein named LvDBP23. This sequence partially matchs one of the probe sequences, but it does not have significant similarity to the gene that was used as a probe. To obtain a full- length mRNA sequence, we decided to carry out 59RACE experiment. The result from 59RACE DNA sequencing further extended 134 nucleotides at 59 end from the primer sequence. The A Novel Shrimp DNA-Binding Protein LvDBP23 mRNA sequence of LvDBP23 consists of 838 nucleotides, including an ORF of 639 nucleotides, 39 UTR of 183 nucleotides with the stop codon (TAA) and polyadenylation signal of CATAAA sequence (Fig. 1). The mRNA sequence was submitted to NCBI GenBank (Accession number: JF742606). The deduced sequence of 212 amino acids from cDNA has a predicted molecular mass of 23.32 kDa. The BLAST result showed the similarity of LvDBP23 to glycine-rich protein of Drosophila melanogaster (Accession No. NP651999) with 44% identity in amino acid sequences. By protein alignment analysis of different glycine- rich proteins, it indicates that the glycine-rich region of LvDBP23 was presented on amino acid position 51–169 (Fig. 2.). In the motif scan bioinformatics software, the domain of Gly-rich is located in amino acid position 46–164 of LvDBP23 protein. The LvDBP23 amino acid sequence was also searched for conserved domain identifications. The Gly-rich domain is predicated to have DNA- binding function. Further online analysis indicates that DNA- binding site of LvDBP23 protein is located on the Gly-rich domain at the amino acid position 94–130. These data suggest that LvDBP23 is a putative DNA-binding protein. Production of LvDBP23 recombinant protein in SF9 insect cell line Compared with bacterial expression system, insect cell expres- sion system is very useful for functional study of eukaryote protein because it can provide post-translational protein modification [7,8]. To test the LvDBP23 mRNA sequence for appropriate translation to a protein, we used SF9 insect cells for producing recombinant protein. GFP and LvDBP23 cDNA were inserted to pIEx-5 expression vector. Successful transfection of the plasmids was confirmed by observing green fluorescent signal with GFP plasmid with phase contract spectroscopy at 0, 24, 48, 72 hr after transfection. LvDBP23 protein was detected by western blot Figure 1. Nucleotide and deduced amino acid sequences of LvDBP23 cDNA. Amino acids are indicated as single capital letters under each triplet codon of the nucleotide sequence. The start codon is the bold underline type and an asterisk (*) indicates the stop codon. The polyadenylation signal (CATAAA) is boxed. doi:10.1371/journal.pone.0019959.g001 PLoS ONE | www.plosone.org 2 May 2011 | Volume 6 | Issue 5 | e19959