| Under the conditions of consistent breeding management and living environment,some of The hybrid F1（Direct cross F1）crossed grass carp（Ctenopharyngodon idellus）and barbel chub（Squaliobarbus cunrriculus）were observed the obvious differences at the size among individuals.This study investigated the phenomenon of segregation of growth traits in hybrid F1.In the study,we used three methods including histology section,real time pcr and race-pcr.The morphological structure of intestinal tissue and the expression level of genes related to growth axis in different growth groups of hybrid F1 were preliminarily analyzed.The structural characteristics and differences of GH and GHR genes in hybrid F1 and parents were compared.The purpose of this study is to explore the factors that affect the growth rate of hybrid F1 in the same environment,which is of great significance to further study the genetic mechanism of the growth difference of hybrid fish.The main research results are as follows:（1）Intestinal tissue characteristics of Direct cross F1 growth difference group:paraffin sections and HE staining were used to observe the intestinal tissue morphology of the growth difference group,the intestinal tissue was composed of mucous layer,submucosa,muscle layer and serous layer in the fast-growing and slow-growing group of F1.The villi of the anterior and middle intestine were long,compact,neatly arranged,the muscular layer was thicker.But the villi of the lower intestine were short,and the villi of the hindgut in the slow-growing group were sparse and scattered.The height of villi and the thickness of myometrium in the fast-growing group were significantly higher than those in the slow-growing group（P<0.05）.（2）Analysis of expression of genes related to growth axis of Direct cross F1:tested the relative expression of GH,GHR,IGF-I,IGF-II,PACAP,SRIF,MSTN-1,MYOG mRNA in release and effect site tissues of two groups of direct cross F1 by used RT-PCR method.The expression of IGF-I and IGF-II in liver of fast-growing group was significantly higher than that of slow growth group（P<0.01）,the expression of SRIF in the pituitary gland of the slow-growing group was significantly higher than that in the fast-growing group（P<0.01）,the expression of GHR and IGF-I in the blood in the fast-growing group was significantly higher than that in the slow-growing group（P<0.01）,and the expression of IGF-I in the muscle in the fast-growing group was significantly higher than that in the slow-growing group（P<0.01）.（3）Cloning and expression of growth hormone and receptor genes in Direct cross F1:the full cDNA sequences of GH（MH046915）and GHR（MH046917）gene in direct cross F1were obtained by using RACE-qPCR method.The complete sequences of GH and GHR cDNA were 1135 bp and 2 660 bp respectively.Their ORFs were 630 bp and 1 815 bp,respectively,encoding 201 amino acids residues and 605 amino acids residues,respectively.There were three and four mRNA unstable signals"attta",with typical Poly A tail,respectively.There were 2 and 6 N-glycosylation sites in GH and GHR amino acid sequences,5 and 6 conserved cysteine amino acid residues,respectively,and GHR amino acid sequences also contained conserved FGEFS motifs,Box1 and Box2.Comparison of nucleic acid and amino acid sequences of direct cross F1 with grass carp and barbel chub showed that GH contains signal peptide and Hormon-1 domain,and GHR was composed of signal peptide,ligand binding domain FN3 domain,transmembrane domain and GHBP domain.The GH amino acid sequence of direct cross F1 was completely consistent with that of barbel chub,and there were only two different sites with grass carp.The GHR amino acid sequences of direct cross F1 and grass carp had 8 difference sites,and there are 12differences between direct cross F1 and barbel chub.The expression of direct cross F1 GH was mainly found in the pituitary gland and the lower level was also detected in the external pituitary tissue.GHR is widely expressed in 12 different tissues and organs. |
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