Cloning And Expression Analysis Of Pancreatic Lipase Gene From Litopenaeus Vannamei And Penaeus Monodon

Fat plays an important role in the formation process of shrimp meat quality and flavor.Pancreatic lipase of lipase family plays an important role in the metabolism of fat inanimaland human.The method of cloning16Sr RNA gene was used to identify that the samplespurchased from Tokyo Station market was the experiment required shrimp Litopenaeusvannamei and Penaeus monodon. As the Marathon kit which choiced in this experiment forthe quality requirements of ds-cDNA, commonly used reference gene beta actin (β-actin)gene was used to detect the quality of ds-cDNA. Using RT-PCR and and RACE-PCR, thefull-length cDNA sequences of pancreatic lipase gene was cloned from Litopenaeusvannamei and Penaeus monodon, and study of pancreatic lipase gene from Litopenaeusvannamei which treated with different salinity. The enzyme method was used in breedingexperiments to measure the activity of lipid–related enzyme with different salinitytreatment. The main results are showed as follow:1) The full length cDNA of Litopenaeus vannamei’s pancreatic lipase (GenBankaccession number of the gene KJ013599) contained1823bp nucleotides, the entire openreading frame (ORF)1494bp codes497amino acids with an estimated molecular mass of54.688kD. The5’-untranslated region (5’-UTR) contained304bp,3’-untranslated region(3’-UTR) contained25bp. The start codon is located from305bp to307bp, and the stopcodon is located from1799to1801bp.2) The full length cDNA of Penaeus monodon pancreatic lipase cDNA full-length,black tiger shrimp pancreatic lipasecontained1976bp nucleotides, the entire open readingframe (ORF)1278bp codes425amino acids with an estimated molecular mass of46.768kD. The5’-untranslated region (5’-UTR) contained535bp, and the3’-untranslated region(3’-UTR) contained163bp. The start codon is located from526bp to538bp, and the stopcodon’s position is from1814bp to1816bp.3) Based on real time PCR technology, the analysis of pancreatic lipase gene fromLitopenaeus vannamei which treated with different salinity in some tissue shows that: thepancreatic lipase gene was expressed both in Litopenaeus vannamei’ muscle and liver afterdifferent salinity treatment, and the expression in liver were higher levels with0.10±0.001-0.085±0.010, where the weak expression in muscle tissue. When salinity at28‰, therewere highest expression both in muscle tissue and liver, while salinity at22‰, the expression was lowest both in muscle tissue and liver. When the salinity at10‰,16‰,34‰and40‰, the expression was no significant difference (P>0.05).4) In Zhanjiang East Island shrimp seed breeding bases, the study of effects onshrimp’s part lipid-related enzyme under different aquaculture water salinity shows that:there is a significant effect on total lipase activity (P <0.05) for Litopenaeus vannamei.When salinity from24‰to30‰and36‰to42‰, the total lipase enzyme activity ofLitopenaeus vannamei was increased with salinity; when salinity from18‰to24‰and30‰to36‰, the total lipase enzyme activity decreased with increase in salinity; in salinity30‰, the total lipase enzyme activity reached maximum. There is’t a significant effect onMDH lipase activity (P <0.05) for Litopenaeus vannamei. When salinity30‰, the MDHlipase enzyme activity reached maximum.