Table 1.
NusG Couples Late after Transcription Initiation
| Strain | lacZ | nutL | Fold Increase of RNA Level (BCM−) | Fold Increase of RNA Level (BCM+) | RT (%) |
|---|---|---|---|---|---|
| MDS42 | wt | – | .25 ± 0.04 | .26 ± 0.03 | 96 ± 19 |
| RSW1225 | SD- | – | .12 ± 0.03 | .56 ± 0.10 | 21 ± 7 |
| RSW1245 | arg(6)—early | – | .13 ± 0.01 | .49 ± 0.02 | 27 ± 2 |
| RSW1276 | arg(6)—late | – | <.001 ± 0.002 | .12 ± 0.003 | <1 |
| RSW1297 | SD- | + | .01 ± 0.006 | .59 ± 0.01 | 2 ± 1 |
Expression of lacZ was induced for 20 min from the lac operon with 1 mM IPTG. Where indicated, Rho-dependent termination was inhibited by adding 100 μg/mL BCM 1 min prior to induction. Readthrough was calculated from the fold increase of lacA RNA compared with ompA RNA in the presence or absence of BCM. RNA levels were measured using qRT-PCR and the fold increase was calculated using the ΔΔCt method (Livak and Schmittgen, 2001). Values are the average of ≥3 independent experiments, each carried out in duplicate. RSW1225 carries two G to A mutations in the lacZ ribosome-binding site. RSW1245 carries an insertion of six rare arginine codons (atg-acc-atg-AGG-AGA-CGA-AGG-AGA-CGA) at the amino terminus of lacZ. RSW1276 contains six rare arginine codons 200 nt distal to the start site of translation. RSW1297 carries an insertion of λnutL immediately 5′ to the mutated ribosome binding site.