The Construction Of The Epider-MIC-Specific Expression Vectors And The Study Of Transgenic Mouse For Thymosin Beta4

As a member of a small actin-binding peptide family, thymosinβ4(Tβ4) could promote hair follicle development and hair growth. In order to understand the role of thymosinβ4on the hair follicle development and hair growth, two epidermic-specific expression vectors were constructed initially. The ability of the promoter has been confirmed by the transfection of Arbas Cashmere goat ear skin fibroblasts. Transgen-ic mice were produced by prokaryotic injection technology, in which4mice were identified as positive transgenic mice by PCR method. In the tails tip of positive transgenic mice, the expression level of thymosinβ4was tested on mRNA levels.1. The construction of epidermic-specific expression vectors pODsRed-KTP and pCDsRed-KTPTwo epidermic-specific expression vectors, pODsRed-KTP and pCDs Red-KTP, have been constructed successfully. In pODsRed-KTP, keratin14promoter (pKRT14) was used to drive thymosinβ4over-expression and Oct-4promoter (pOct-4) drived the red fluorescent protein. As the control, pCDsRed-KTP used pKRT14as the pro-moter of thymosinβ4and in which the red fluorescent protein was drived by CMV promoter (pCMV).The pKRT14, an epidermic-specific promoter, could drive thymosinβ4express-ing at epidermic cells with the result that pODsRed-KTP and pCDsRed-KTP had the feature of epidermic-specific expression. In pODsRed-KTP, the red fluorescent pro-tein drived by pOct-4expressed in certain kinds of stem cells so as to display the stem cells’directions. In pCDsRed-KTP, the red fluorescent protein drived by pCMV expressed in general cell lines. The main function of pCDsRed-KTP was as the control in the next experiment.2. The verification of pKRT14by transfecting to Arbas cashmere goat ear skin flbroblastsEpidermic-specific expression vectors, pODsRed-KTP and pCDsRed-KTP were transfected to Arbas cashmere goat ear skin flbroblasts. Observed the red fluorescent of cells with pCDsRed-KTP, the transient transfection efficiency of cells with pODsRed-KTP has been known, about thirty-one percents. In stable thymosinβ4ex-pressed cells, the expression level of thymosinP4was testing on mRNA level. Com-pared with untransformed cells, the expression level of thymosinβ4was up-regulated4.51times in the transfected cells. Results showed that pKRT14was efficient to drive thymosinβ4expression. 3. The production and identification of transgenic mice for thymosinβ4Fifty mice were produced by prokaryotic injection technology, in which4mice were identified as positive transgenic mice by PCR method, including1male and3females, the rates of positive transgenic mice for thymosinβ4was eighth percent. In the tails tip of positive transgenic mice, the expression level of thymosinβ4was tested on mRNA and protein levels. Compared with the control, the thymosinP4expression mRNA level in transgenic mice ranged from1.32-2.05times. Positive transgenic mice were produced and identified so that to lay the foundation for further characterization of thymosinβ4functions and other related researches.