Full length cDNA cloning and tissue expression of major histocompatibility complex (MHC) class Ⅰfrom blunt snout bream (Megalobrama amblycephala)

Abstract:

RACE (Rapid Amplification of cDNA Ends) technique was used to clone “Pujiang No.1” blunt snout bream (Megalobrama amblycephala) MHC class Ⅰgene (Meam-MHC I), and total three full-length cDNA sequences in the F₀ and F₈ generations were obtained by cloning and sequencing method, respectively. The sequences were 2 040-2 079 bp in length, including an 87-102 bp 5′-UTR, a 1 035-1 044 bp coding region, containing signal region, three (α1, α2, α3) domains, a transmembrane and cytoplasmic region (TM/CY), and 911-946 bp 3′-UTR. The coding region had 344-347 amino acids. Within F₈ generation, the nucleotide/amino acid sequences similarities were as high as 89.0%/93.0%. But the values were relatively as low as 75%-77% /71% between generations, indicating obviously sequence polymorphism. The Meam-MHC I exhibited a classical 3D molecular structure, and compared to the PBR region of HLA-A2,the major differences were mainly in the five regions (I-V). It was revealed in the neighbor-joining tree that the blunt snout bream had the closest relationship with grass carp, far from salmon and trout, and still farther from, chicken and human. Semi-quantitative RT-PCR analysis demonstrated that Meam-MHC I transcript was detected at high level in gill, head kidney and blood, moderate in liver and spleen, and low in trunk kidney, gut and muscle.