Study Expression Pattern Of Gonad Related Genes From Large Yellow Croaker Larimichthys Crocea

WD40 repeat proteins,contain varying numbers of conserved WD repeats domain of 40 amino acids and play an important role in cell cycle.In the present study,we cloned full-length cDNA sequences of the genes wdr55,wdr73,wdr91,dcaf17 from the important commercial fishes in china-large yellow croaker(Larimichthys crocea)by the method of rapid-amplification of cDNA ends(RACE),(that were named as Lc-wdr55,Lc-wdr73,Lc-wdr91,Lc-dcaf17 respecctively).Moreover,using quantitative real-time PCR to detect their expression in different tissues of large yellow croaker,the same as to various developmental stage of gonads.The results as follows:1.The full-length cDNA sequence of Lc-wdr55 is 1,849 bp,including an open reading frame(ORF)of 1,233 bp which encoded 410 amino acids and had seven conserved domain and EWD box of WD40 family.The expression of Lc-wdr55 in the ovary and testis was significantly higher than other tissues(P <0.05),and that the expression level in testis is higher than the ovary.In different stage of gonad,Lc-wdr55 expression was the highest level at mature stage,then proliferative phase,voiding stage was lowest level(P<0.05);which show the significant correlation with gonad development and mature.2.The whole cDNA sequence of Lc-wdr73 is 1,706 bp in length including an ORF of 1,406 bp which encoded 381 amino acids.5’UTR is 51 bp,3’UTR is 509 bp,protein molecular mass of 41.381 kDa,an isoelectric point of 5.15.Lc-wdr73 was also expressed in multiple tissues and had highest level in ovary and testes.The expression of Lc-wdr73 in gonad increased gradually as the gonad development and mature,and reached the highest level at mature stage,and then it decreased sharply to the lowest level at the voiding stage(P<0.05).Meanwhile,the expression of Lc-wdr73 was with the significant higher expression in testis than in ovaries at the same stage except voiding stage during the whole reproductive cycle.We inferred that Lc-wdr73 contributed to the developmental processes of gonad and germ cell maturation,and the Lc-wdr73 contribute much more to ovary than testis.3.Lc-wdr91 full-length including 2,711 bp,ORF 2,259 bp encoding a predicted 752 amino acid.The molecular weight of 84.430 kDa,isoelectric point 6.32.Five conserved WD40 repeat motif existed in Lc-wdr91.Lc-wdr91 mRNA expressed wildly in Juvenile,young and adult fish tissues and development cycle(proliferative stage,growing stage and mature stage voiding tage)of L.crocea,and the testis had most expression,the second was ovary(P<0.05).The above results show that the Lc-wdr91 have correlated with gonad development cycle,However,Lc-wdr91 are getting far more involved in testes.4.Lc-dcaf17 gene,the full-length cDNA is 1,733 bp,containing an open reading frame(ORF)of 1,530 bp which encoded a deduced protein of 509 amino acids.The molecular weight of the deduced protein was 89.281 kDa and the isoelectric point was 5.05.The qPCR demonstrated that Lc-dcaf17 was expressed in all tested tissues of Juvenile,young and adult fish with the highest expression level in gonads,predominadely expressed in ovaries and secondly expressed in testes.With the gonad development,Lc-dcaf17 expression exhibited a increasing trend and peaked in maturation stage and ebbed in void phase(P<0.05).These results suggested that Lcdcaf17 might be involved in gonad development in L.crocea.