通过RNA干扰降低肌肉生长抑制素基因,增加鸡的体重.pptxVIP

1;Introduction;4.Increased expression ofmyostatin was also observed in humans with chronic illness, HIVinfection and early aging due to muscle atrophy. 5.The limitingof function of negative regulator of any trait can be achieved by anumber of techniques of which shRNA based knock down approachwould be a promising one. 6.In this study, chicken has been used asmodel organism to estimate the effect of shRNA on phenotypes.;AnimalsControl broiler birds maintained at the Institute farm Slaughter of birds was performed fol-lowing standard protocol of cervical dislocation MSTN gene (Accession No. AF346599) Fertile eggs of control broiler chicken line werecollected from the Institute farm, scrambled and control birds using Trizol accord-ing to manufacturer’s instruction (Sigma). The analysis kit (Jeev Diagnos-tics Pvt. Ltd, India);Experiment process;Fig. 1. Secondary structures of antisense strands of shRNAs for myostatin gene predicted at the site;Fig. 2. Location of ink in the yolk inoculated through 26 gauge needle from narrow end of egg. A length of 2 cm of the needle inserted into the egg and spot shows the locationof ink in the yolk of the egg. ;;;Fig. 3. PCR product in screening of positive knock down birds amplified from DESTvector.;Fig. 4. Southern blot of genomic DNA digested with PstI enzymecollected fromknock down chicken containing shRNA molecules Mol1 to Mol5 including scrambledand control group.;Fig. 5. The mRNA expression level in knock down birds with 5 shRNA molecules, birds of scrambled shRNA molecule and control group revealed differential expressionamong groups where birds of molecule4 had the lowest expression of MSTN mRNA;Fig. 6. Western blot of myostatin expressed in breast muscle tissue collected from knock down chicken containing shRNA molecules Mol1 to Mol5 including scrambled andcontrol group.;Fig. 7. In-situ histochemistry of breast muscle of knock down chicken carrying shRNA 4 molecule revealing blot of MSTN protein inside the myoblast cells. ;