cDNA cloning and primary structure analysis of invariant chain in Chinese Pengze crucian carp

Abstract

Invariant chain (Ii) plays an important role in MHC class II molecules assembly and exogenous peptide presentation in vertebrates. Although mammalian Ii has been extensively studied, less attention is paid to its fish counterpart. In this study, in order to understand the structure and biological function of Chinese Pengze crucian carp (Carassius auratus) Ii, we first cloned Ii gene (Caau-Ii) using RT PCR and rapid amplification of cDNA end techniques. Firstly, a conserved DNA fragment was amplified using a pair of degenerate primers from fish’s head kidney and spleen total RNA. Then specific primers for RACE were designed to amplify the 5' and 3' ends of Caau-Ii. Finally, the full-length cDNA of Caau-Ii was amplified, sequenced and analyzed and the results showed that it was 1063 bp in length and contained an open reading frame of 696 bp encoding Caau-Ii precursor protein. Amino acid sequence alignment showed that (1) Caau-Ii had high homology with that of carp and zebrafish (identify >70%) and low homology with mammalian Ii (30% < identity <40%); (2) Caau-Ii had similar domains seen in mammalian Ii, including transmembrane domain, CLIP (class II-associated Ii-derived peptide) and TRIM (trimerization) domain.

Key words: Invariant chain, Carassius auratus, RACE, cloning, domain analysis.