Highly variable polymorphism of the alpha-amylase gene family in Litopenaeus vannamei (Crustacea Decapoda).

Alpha-amylase from the tropical shrimp Litopenaeus vannamei presents a high degree of polymorphism and at least eight different electromorphs are detected by electrophoresis. Based on nucleotide sequences, three cDNAs have been previously characterized. In this paper we report on the organization and the evolution of corresponding alpha-amylase genes, determined after PCR amplification. Three AMY genes have been characterized, spanning over 3.3 kb and encoding mature proteins of 495 amino acids (aa), which are all expressed in the digestive gland. The existence of nine short introns, ranging from 86 to 454 bp, located at the same positions for each of the different genes, and presenting no similarity between them, is reported. Between 11 and 15% of changes are observed in the coding aa sequences of genes II and III compared to the gene I sequence respectively. One 5' putative promoter sequence has been sequenced and shows no classical TATA box upstream to the coding sequence. Based on the intron size difference, a single PCR (producing the S-R fragments) allows the separation of a partial gene I (750 bp), corresponding to cDNA 20, from the others (650-680 bp). Sequencing different S-R PCR fragments from one shrimp shows at least eight different haplotypes. A complex microsatellite repeat is present in intron 6 of gene II. Using size and sequence differences in this repeated portion, it is possible to characterize two gene subfamilies (IIa and IIb) encoding previously described cDNAs 28 and 37, respectively. For the gene II family, two to four alleles are present in one shrimp corresponding to these two genes. Within the Panama natural population, 35 different alleles are shown at this locus. Regarding alpha-amylase gene structure in the shrimp, many recombinants are present from a set of individuals and constitute an important mechanism of evolution of alpha-amylase function.