1. Resuspend cDNA in 27 pL ddH2O. An example reaction is the following:
27 pL cDNA;
5 pL 10X ligase buffer;
6 pL 32P-labeled linkers—0.8 pg;
3 pL unlabeled linkers—3 pg;
2 pL T4 RNA ligase—12-18 U (see Note 6). Incubate for 1 h at room temperature then 14°C overnight.
2. Heat-kill the enzyme at 80°C for 15 min, and then cool on ice.
3. Remove 1 pL of the ligation mix to test for ligation (see Note 7).
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