title: Genomic structure of the α-amylase gene in the pearl oyster Pinctada fucata and its expression in response to salinity and food concentration.
abstract: Amylase is one of the most important digestive enzymes for phytophagous animals. In this study, the cDNA, genomic DNA, and promoter region of the α-amylase gene of the pearl oyster Pinctada fucata were cloned by using reverse transcription-polymerase chain reaction (RT-PCR), rapid amplification of cDNA ends, and genome-walking methods. The full-length cDNA sequence was 1704bp long and consisted of a 5'-untranslated region of 17bp, a 3'-untranslated region of 118bp, and a 1569-bp open reading frame encoding a 522-aa polypeptide with a 20-aa signal peptide. Sequence alignment revealed that P. fucata α-amylase (Pfamy) shared the highest identity (91.6%) with Pinctada maxima. The phylogenetic tree showed that it was closely related to P. maxima, based on the amino acid sequences. The genomic DNA was 10850bp and contained nine exons, eight introns, and a promoter region of 3932bp. Several transcriptional factors such as GATA-1, AP-1, and SP1 were predicted in the promoter region. Quantitative RT-PCR assay indicated that the relative expression level of Pfamy was significantly higher in the digestive gland than in other tissues (gonad, gills, muscle, and mantle) (P<0.001). The expression level at salinity 27‰ was significantly higher than that at other salinities (P<0.05). Expression reached a minimum when the algal food concentration was 16×10(4)cells/mL, which was significantly lower than the level observed at 8×10(4)cells/mL and 20×10(4) cells/mL (P<0.05). Our findings provide a genetic basis for further research on Pfamy activity and will facilitate studies on the growth mechanisms and genetic improvement of the pearl oyster P. fucata.
authors.affiliation: Key Laboratory of South China Sea Fishery Resources Exploitation & Utilization of Ministry of Agriculture, Guangzhou 510300, China; South China Sea Resource Exploitation and Protection Collaborative Innovation Center (SCS-REPIC), Guangzhou 510300, China; South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou 510300, China.
authors.forename: Guiju
authors.initials: G
authors.lastname: Huang
authors.affiliation: Key Laboratory of South China Sea Fishery Resources Exploitation & Utilization of Ministry of Agriculture, Guangzhou 510300, China; South China Sea Resource Exploitation and Protection Collaborative Innovation Center (SCS-REPIC), Guangzhou 510300, China; South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou 510300, China.
authors.forename: Yihui
authors.initials: Y
authors.lastname: Guo
authors.affiliation: Key Laboratory of South China Sea Fishery Resources Exploitation & Utilization of Ministry of Agriculture, Guangzhou 510300, China; South China Sea Resource Exploitation and Protection Collaborative Innovation Center (SCS-REPIC), Guangzhou 510300, China; South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou 510300, China.
authors.forename: Lu
authors.initials: L
authors.lastname: Li
authors.affiliation: Key Laboratory of South China Sea Fishery Resources Exploitation & Utilization of Ministry of Agriculture, Guangzhou 510300, China; South China Sea Resource Exploitation and Protection Collaborative Innovation Center (SCS-REPIC), Guangzhou 510300, China; South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou 510300, China.
authors.forename: Sigang
authors.initials: S
authors.lastname: Fan
authors.affiliation: Key Laboratory of Marine Bio-resources Sustainable Utilization, Chinese Academy of Sciences, Guangzhou 510300, China.
authors.forename: Ziniu
authors.initials: Z
authors.lastname: Yu
authors.affiliation: Key Laboratory of South China Sea Fishery Resources Exploitation & Utilization of Ministry of Agriculture, Guangzhou 510300, China; South China Sea Resource Exploitation and Protection Collaborative Innovation Center (SCS-REPIC), Guangzhou 510300, China; South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou 510300, China. Electronic address: pearlydh@163.com.
authors.forename: Dahui
authors.initials: D
authors.lastname: Yu
mesh_terms: D000818:Animals; D003001:Cloning, Molecular; D010802:Phylogeny; D049911:Pinctada / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000502:physiology; D054712:Salinity; D059467:Transcriptome; D000516:alpha-Amylases / Q000737:chemistry / Q000235:genetics*
chemical_list: D000516:alpha-Amylases
keywords: Algal concentration; Bivalve; Digestive enzyme; Environmental factors; Tissue expression; cDNA cloning