反応式
[As](C)(C)(=O)[O-].C(S)[C@@H](O)[C@H](O)CS.[P:14]([O:26][CH2:27][C@H:28]1[O:32][C@@H:31]([N:33]2[CH:40]=[CH:39][C:37]([NH2:38])=[N:36][C:34]2=[O:35])[CH2:30][C@@H:29]1[OH:41])([O:17]P(OP(O)(O)=O)(O)=O)(=[O:16])[OH:15].C(N(CC(O)=O)CC(O)=O)CN(CC(O)=O)CC(O)=O>O>[CH2:30]1[C@H:31]([N:33]2[C:34](=[O:35])[N:36]=[C:37]([NH2:38])[CH:39]=[CH:40]2)[O:32][C@H:28]([CH2:27][O:26][P:14]([OH:16])([OH:17])=[O:15])[C@H:29]1[OH:41]参加物
実験手順
Double-stranded cDNA was homopolymer tailed essentially by the method of R. Roychoudhury and R. Wu, Meth. Enz., 65, pp. 43-62 (1980). The following were added together, in order: 3 μl dCTP32 (>800 Ci/mmol), 20 μl 10 X cacodylate buffer (1.4 mM potassium cacodylate, 0.3M Tris-hydrochloride, pH 7.6), 4 μl 5 mM DTT, 3 μl 10 mM dCTP, 2 μl 100 mM CoCl2, 12 units terminal deoxynucleotide transferase (PL Biochemicals; at least 8 units/pmol 3' end) in 1.5 μl, H2O to 200 μl final volume. The reaction was carried out at 12° for 1.5 min then stopped by adding 10 μl 0.5M EDTA This reaction resulted in an average of 15 dCMP residues added per 3' end of dsDNA, the optimal number for annealing and transformation (Peacock et al., Bioch. Biophys. Acta, 655, pp. 243-50 (1981)
後処理
- additionThe following were added together, in order
出典と根拠
- ORDレコード
- ord-0c084c41e25d412587e8f744f87f5241
- 出典データセット
- uspto-grants-1993_04
- 特許番号
- US05202430
関連ノート
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