Table 2.
Mutations generated in the cro gene during in vitro gap-filling replication
| Mutation type | No. of mutations
|
||
|---|---|---|---|
| Pol III† | Pol V‡ | No polymerase§ | |
| Base substitution | 24 | 72 | 42 |
| Frameshift | 20 | 20 | 0 |
| Other¶ | 27 | 9 | 0 |
| All mutants | 71 | 101 | 42 |
| Transition | 13 | 23 | 27 |
| A → G | 0 | 0 | 1 |
| C → T | 11 | 5 | 24 |
| G → A | 2 | 2 | 1 |
| T → C | 0 | 16 | 1 |
| Transversion | 11 | 49 | 15 |
| A → C | 2 | 7 | 2 |
| A → T | 0 | 18 | 0 |
| C → A | 0 | 4 | 0 |
| C → G | 0 | 1 | 2 |
| G → C | 5 | 5 | 8 |
| G → T | 2 | 1 | 3 |
| T → A | 2 | 10 | 0 |
| T → G | 0 | 3 | 0 |
Gap-filling replication reactions were performed with the indicated DNA polymerases, after which the DNA products were introduced into an E. coli indicator strain and plated on lactose-EMB plates. Plasmids were extracted from dark-red mutant colonies, and the sequence of their cro gene was determined by DNA sequence analysis. The details are presented in Materials and Methods.
†Replication was performed with pol III holoenzyme.
‡Replication with pol V, in the presence of UmuD′, RecA, and SSB.
§Nonreplicated gapped plasmid was used to transform the indicator strain.
¶ Other mutations include big deletions and insertions as well as complex mutations.