Cloning And MRNA Expression Of Candidate Genes For Growth Performance Of Macrobrachium Rosenbergii

The giant freshwater prawn(GFP),Macrobrachium rosenbergii,is the main cultured freshwater crustacean species in China.Curre ntly,there exits few problems such as uneven specifications,individ ual miniaturization,low yield,etc.,which seriously limited the high-quality and efficient development of GFP aquaculture industry.The refore,the aim of this study was to do research of candidate genes for growth performance of GFP,and to unravel the expression patt erns.Thus,the results will provide a theoretical foundation for furt her exploring the regulatory mechanism of candidate genes during g rowth and development..Based on the population imported from Bangladesh,20 full-sib families were constructed.And their growth performance was com pared,thus the fast-growing group(FG)and the slow-growing grou p(SG)were selected.Then,eight candidate genes related to growth performance including Trypsin(TRY),Amylase(AMY),Cathepsin(CaTL),Myostatin(MSTN),Retinoid X receptor(RXR),Ras-related nuclear protein(Ran),Heat shock protein 90(HSP90)and Ecdyson e receptor(EcR)were cloned and analyzed by bioinformatics;Addti onally,real-time quantitative PCR(RT-PCR)was used to study the gene expression.Hence,the results were obtained as follows:1.Based on GFP population imported from Bangladesh,20 ful 1-sib families were constructed.The results for comparison of growt h performance showed that the growth rates of the 20 families wer e significantly different.Among them,family 19 had the highest bo dy weight,and family 18 had the longest body length.Depend on the above results,five fast-growing families(family 6,11,12,18,19)and five slow-growing families(family 1,3,9,15,17)were se lected out.Further comparative analysis displayed that the body wei ght(BW),lbody length(BL),cephalothorax length(CL),abdominal length(AL),cephalothorax width(CW)and abdominal width(AW).of FG were significantly higher than SG(P<0.01).2.The coding regions of TRY,AMY,CaTL,MSTN,RXR,Ran,HSP90 and EcR of GFP were successfully cloned from hepatopanc reas.The full length of CDS for each gene was 801 bp,2120 bp,1029 bp,1380 bp.1346 bp,648 bp,2181 bp and 1716 bp;the nu mber of amino acids was 266,706,342,459,448,215,726 and 571;The results of amino acid sequence homology alignment showe d that there were 1(c.C589T induced p.Phe298Leu),1(c.G246A in duced p.Tyr84Cys),2(c.T299C induced p.Ile100Thr and c.A503G i nduced p.Asn168Asp)and 1(C.A897G induced p.Ile 298Val)alterat ions in the TRY,AMY,CaTL and EcR of GFP,respectively.There were 9 alterations in the amino acid sequence of HSP90 gene.The amino acid sequence of Ran gene was corresponding to that provid ed by NCBI.And a total of 15 and 21 amino acids of MSTN and RXR were found to be different from those of Macrobrachium nip ponense.3.Using RT-PCR technology,eight tissues consist of gonad,he patopancreas,muscle,gill,stomach,heart,intestine and abdominal n erve of GFP(? and ?)were used to detect the expression profile s of eight candidate genes.It was demonstrated that the expression of TRY and AMY genes were highest in hepatopancreas(? and ?),and the lowest in muscle;the expression of CaTL gene was highe st in hepatopancreas(?)and gill(?),lowest in abdominal nerve.T he highest mRNA expression levels of MSTN gene in the heart(?and ?),and the lowest in the stomach(?)and muscle(?).The RXR gene had the highest expression in gill(?)and ovary(?),and the lowest expression in intestine(?)and hepatopancreas(?);The Ran gene is highly expressed in gonad tissue,and lower expr essed in intestine(?)and stomach(?);The expression of HSP90 gene was highest in hepatopancreas(?)and heart tissue(?),low est expression in muscle(? and ?);The mRNA expression of Ec R gene was highest in gill(?)and ovarian tissue(?),and it has a low expression level in muscle(?)and abdominal nerve(?).4.In order to further explore the expression patterns of eight c andidate genes,the muscle tissues of 8 individuals in FG and SG were collected for RT-PCR experiments.The results revealed that th e mRNA expressions of TRY,AMY,CaTL,HSP90 and EcR genes i n FG were significantly higher than those in SG(P<0.01 or P<0.05).The mRNA expression level of MSTN in FG was significantly 1 ower than that in SG(P<0.01).There was no significant difference in the expression of Ran and RXR gene between the two groups(P>0.05).