Studies On Different Of Proteins In Hair Follicles Of The Skin Tissue Of Fine-wool Sheep With Different Fiber Diameter Using Two-dimensional Electrophoresis And Mass Spectrometry

This Experiment chosen the different diameters of follicles which taken from thefine-wool sheep’s skin tissue as the test material, use different extraction methods, differentpH values of the IPG adhesive strip and different amount of sample quantity. Aim to exploretwo dimensional electrophoresis system of fine wool sheep skin tissue, and built up2-DE gelchromatogram for fine-wool sheep’s skin tissue, afterward, find out the differential protein bymass spectrometry, onwards, identify its peptide mass fingerprinting and alignment them inthe database, then combine the Bioinformatics to analyse the internal relationship betweenthese proteins and the diameters of wool fabrics. The main obtained the following results:1. Through comparison of the TCA precipitation method, the phenol extraction and theTCA/acetone method, find out the suitable extraction of the total protein from the fine-woolsheep’s follicles tissue.2. The IPG adhesive strip was picked PH3-10to separate the follicles’ total protein fromskin tissue, and focus on exploring the density range of protein spots. In this step, in the wayof separating the follicles’ total protein spot range more distinguished on the2-DE map, usedIPG adhesive strip is in the range of PH4-7and the length of18cm IPG linear tape. This helpsthe total protein which extract from follicles distribute properly. After compare the80μL,100μL and120μL on each sample of strip respectively, then get the clearest map is100uLstrip, which IPG adhesive strip is linear tape is18cm, pH4-7, get the good quality oftwo-dimensional gel electrophoresis.3. Use2year old Super thin and thin wool hair follicle’s protein analyse on the2-DE gelchromatogram, test out the difference point7,30,31,34,35comes out reduce, the rest ofprotein all rise up. We can see from the figure above compared with super thin, four thin hairfollicles all reduce. In the reducing regulation proteins point7comes out the differentialprotein. After analyse2-DE gel chromatogram from4years old super thin and thin wool hairfollicle which detected expressed more than2times38differentially protein. In addition, thedifferential protein comes out down regulation which is36,57and58, the rest differentialprotein were up regulated.36,57and58differential protein all comes from the thin wool hair.Comparison of the different diameters of hair follicles’ protein, we got73mass spectraand73sets of peptide mass fingerprint result, then search in the database of the NCBI, all these73sets of protein can be expressed the different protein. Compare the different age ofsuper thin wool hair follicles’ protein, we got33mass spectra and33sets of peptide massfingerprint results. In this results, there are12protein groups with different fiber diameter ofwool hair protein groups repeated. In this experiment, according to the main function, the94proteins is divided into8categories.