Establishment and biological characteristics of Ujumqin sheep fibroblast line.

A Ujumqin sheep ear marginal tissue (USEM) fibroblast line, frozen in 147 cryovials with 4 x 10(6) cell each, was successfully established from 33 Ujumqin sheep ear marginal tissues using explant culture and cryopreservation techniques. The cells were morphologically consistent with fibroblasts. The growth curve was typical S-shape and the cell population passed through a lag phase, a logarithmic phase and a plateau phase. The population doubling time (PDT) was approximately 72 h. Tests for bacteria, fungi, viruses and mycoplasma were all negative. Isoenzyme polymorphism indicated that the genetic characteristics of the cell line were stable in vitro. Karyotyping analysis indicated that the chromosome number of a normal cell was of 2n = 54 and 95.4% of the entire population was diploid. The transfection efficiencies of six fluorescent proteins (pEGFP-N3, pEGFP-C1, pDsRed-N1, pEYFP-N1, pECFP-N1 and pECFP-mito) optimal at 48 h were from 18.5% to 30.1%. The cell line met all criteria from the American Type Culture Collection (ATCC). Not only has the germline of this important sheep breed been preserved at the cell level, but also valuable material had been provided for genome, postgenome and somatic cloning research. Moreover, the establishment of this technical platform may provide both technical and theoretical support for storing the genetic resources of other animals and poultry at the cell level.