Molecular cloning and expression and phylogenetic trees’ construction of CYR61 gene in common carp

Abstract:

CYR61,CTGF and NOV constitute the CCN family. We used BLASTN program at the NCBI to identify near matches in the common carp (Cyprinus carpio) genome database to each zebrafish (Danio rerio) CYR61 .CYR61 A, CYR61 B, and CYR61 C were identified respectively. ORF of the 3 duplicates of CYR61 were cloned and sequenced in common carp. CYR61 A was 1 158 bp encoding polypeptides of 385 amino acids with a calculated Mw of 42.55 ku and PI of 7.83. CYR61 B was 1 053 bp. CYR61 C was 1 107 bp encoding polypeptides of 368 amino acids with a calculated Mw of 40.83 ku and PI of 8.54. CYR61 A and CYR61 C have 5 exons and 4 introns. CYR61 A, CYR61 B, and CYR61 C all contained 4 conservative domains in common carp. Phylogenetic relationship analysis indicated that CYR61 genes of common carp have higher homology with other species’ CYR61 genes. Differentiation time of CYR61 A, CYR61 B and CYR61 C were earlier than the divergence time between common carp and zebrafish by using MEGA 5.0 software and divergence time formula analysis. RTPCR analysis revealed CYR61 C to be expressed in most common carp tissues, with the principal expression in ovary and weakest expression in heart. CYR61 A was predominantly expressed in testicle and not detected in kidney and body kidney. CYR61 B was highly observed in testicle and brain. Quantitative realtime PCR were used to analyse the expression of CYR61 in common carp development, the result indicated that CYR61 transcriptions expressed highly at the initial stage and lowest at 18 hpf or 24 hpf and then increased before 3d， but decreased at 6 d.