| First Author | Czank A | Year | 1991 |
| Journal | Gene | Volume | 109 |
| Issue | 2 | Pages | 259-63 |
| PubMed ID | 1662657 | Mgi Jnum | J:729 |
| Mgi Id | MGI:49263 | Doi | 10.1016/0378-1119(91)90618-l |
| Citation | Czank A, et al. (1991) Expression in mammalian cells of a cloned gene encoding murine DNA methyltransferase. Gene 109(2):259-63 |
| abstractText | Mammalian DNA cytosine-5-methyltransferase (MTase, EC 2.1.1.37) is an essential component for establishing and maintaining cell-type specific methylation patterns in the genome. The cDNA for the murine enzyme was previously cloned in segments. We have reconstructed the entire gene, encoding a protein of 1517 amino acids, from a set of overlapping cDNA clones. We report the assembly of two expression constructs in bacterial/mammalian shuttle vectors. Transcription in the first construct (pEMT) is driven by the cytomegalovirus enhancer/promoter and encodes a fusion protein with 15 additional aa at the N terminus, while the second construct (pJMT) is driven by the simian virus 40 early promoter/enhancer upstream from the natural ATG codon. Immunofluorescence microscopy and immunoblot analysis have shown that both constructs direct the synthesis of MTase in COS-1 cells. Enzyme activity in whole-cell lysates of transfected COS-1 cells transfected with pEMT and pJMT are on average tenfold and fivefold higher than in controls, respectively. The specific activities of the recombinant and endogenous mouse-cell enzyme are similar. These expression constructs will be of use in studies of DNA methylation in mammals. |
