Cloing And Expression Analysis Of Related Genes PdGig2 And PdFrr2 During Caudal Fin Regeneration In Paramisgurnus Dabryanus

Mature individuals can reconstruct the lost tissues based on the original organization after damage,this phenomenon is called regeneration.All living organisms will make the corresponding physiological reaction in response to tissue damage and they also are closely related to the occurrence of disease,so it is of great importance to study the mechanism of regeneration process.Mammals regeneration ability is very limited,amphibians represented by the salamanders and bony fish represented by zebrafish can complete the reconstruction of the structure and function by epimorphosis after cutting appendages and fins.But since slow amphibians breeding,hard to raise in lab,lack of corresponding genetic operators,it can’t be the ideal research animals.Fin can complete recovery after cut off the original structure,and is relatively simple,easy to get,don’t harm the body after injury.more significant,bony fish fin share similar early development process with tetrapods involved in human,but both have vast differences in regeneration,so fin can be a good system to analytical Molecular regeneration mechanism.This study first get two great differentially expressed ESTs in fin regeneration from the established SSH c DNA library,on the basis of the two ESTs,adopt the method of RACE for amplification,get two full length c DNA sequences.Through Blast,one of the genes is thought to be Gig2,and another can’t find any highly homologous,we considered it to be a new gene named Frr2 gene.Then use DNAMAN MEGA6.0,sigmaplot software to analyze biological function of two genes,and detect the expression in different tissues.Gig2 gene is grass carp hemorrhagic virus gene induced gene 2,firstly identified from UV-inactivated GCHV treated Carassius auratus blastulae embryonic(CAB)cells.It plays an important role in fish interferon antiviral response,and can adjust regeneration could be a new discovery.c DNA full sequence is 712 bp,encoding 156 amino acids,relative Molecular mass is 17.324 KDa,isoelectric point(p I)8.69.46 polar amino acid,71 non-polar hydrophobic amino acids,18 acidic amino acids,21 basic amino acid.Protein secondary structure found alpha helix accounted for 23.72%,extending chain(27.56%),beta angle is occupied 16.03%,32.69% random coil.Semi-quantitative and fluorescence quantitative detection,the author found out that the 4 d after regeneration is at their peak levels,highest expressed in the membrane phase during embryonic development,also found that expressed in testis tissue is also the biggest.According to Gig2 expression pattern for the first time,we found the genes involved in the process of fin regeneration,it may play a regulatory role in the process of fin structure formation.Cognate degrees found Gig2 gene has relatively low homologous,which attract scientists great interest in their evolutionary relationships.Frr2(fin regeneration related gene2)gene is an unknown,it is a gene associated with fin regeneration.c DNA is 1036 bp,encoding 88 amino acids,secondary structure found alpha helix is occupied 46.59%,beta Angle is occupied 9.09%,random coil accounted for 32.95%,extending chain accounted for 11.36%.Semi-quantitative and fluorescent quantitative detection found in different periods(0 dpa,2 dpa,3 dpa,4 dpa,5dpa,6 dpa)in regeneration,different organs(liver,heart,brain,testis,kidney,ovary),gene expression level in different embryo developmental periods(blastocysts,gastrula,neural embryo,tail bud stage,the membrane phase),found that it is highest expression level in 4dpa,heart,gastrula period.Frozen section in situ hybridization technique apply in different periods in regeneration,found expression in osteoblast,strongest signal in 4dpa.We also found expression in osteoblast,we hypothesized that Frr2 gene might be involved in osteoblast proliferation and differentiation in the process,related to the formation of the fin ray.This two genes change in expression in caudal fin regeneration and reach to high level in later regeneration,it is predicted the genes play a certain role in the process of regeneration,They may control the regeneration process of the formation of the fin structureit,it still remains to be further study about their more functions.