## 2 Result and Discussions
### Influence of P-Expression on Plasmid Retention.  
The _in_ability of plasmids expressing \(P\) to form stable transformants is used as a measure of \"P-lethality\", based on an assumption that P made from the transforming plasmid kills the transformed  
Figure 2: Plaque type / formation by \(\lambda cI72\) on 594[pcIpR-\(P\)-timm] and 594 host cells. The ability of \(\lambda cI72\) to form plaques on 594[pcIpR-\(P\)-timm] host cells at 35-42 \({}^{\circ}\)C is attributed to loss, or considerable reduction, in plasmid copy number per cell. The \(imm\lambda\) interference phenotype (compare EOP, Table 5, 25 and 30 \({}^{\circ}\)C _vs._ 35 \({}^{\circ}\)C) is dependent upon CI[Ts] repressor expressed from the plasmid. The cellular loss of \(imm\lambda\) interference correlates with plasmid loss and the observed increase in plaque size between 35 and 42 \({}^{\circ}\)C; whereas, the plaque size was essentially constant between 30 to 42 \({}^{\circ}\)C when \(\lambda cI72\) was plated on 594 cells without the plasmid. The photos of the individual plaques shown in the top row were taken through the lens of a stereo microscope, from agar overlay plates (middle row) incubated at the indicated temperatures.  
cells. The influence of \(P\) expression from pclpR-_P_-timm in 594 _dnaB\({}^{+}\)_ cells was examined for plasmid replication / maintenance / stability (Figure 3). Plasmid loss was observed in cultures of 594[pclpR-_P_-timm] grown at 34 or 35 \({}^{\circ}\)C (in comparison to cultures grown at 30 \({}^{\circ}\)C), but curing was extensive in cell cultures grown at or above 36 \({}^{\circ}\)C (Figure 3B). Essentially identical results were seen for 594 _clpP_[pclpR-_P_-timm] cultures (Figure 3B) showing that inactivation of the _clpP_ protease, which participates along with ClpX in degrading \(\lambda\) O protein [39,40], did not significantly influence P-dependent plasmid loss relative to the _clpP\({}^{+}\)_ strain 594; the ClpP protease defect did not amplify plasmid loss, which is expected if its absence increased the P concentration per cell. Secondary cultures prepared from 594 _grpD55_[pclpR-_P_-timm], or 594[_pclpR-_P_\({}^{\text{A}76}\)-timm] cells retained their plasmids, even when grown at 42 \({}^{\circ}\)C for at least seven doublings while constitutively expressing \(P\) (Figure 3C). The _grpD55_ allele blocks \(\lambda\) vegetative growth (Table 2) and permits cell survival (at some level) in the presence of constitutive \(P\) expression at 42 \({}^{\circ}\)C, suggesting that allelic variations in Dna_can_ protect cells from \(P\) expression and its consequent negative influence on cell metabolism. This result is in disagreement with the hypothesis made by [PERSON] _et al._[24], which in essence states that allelic variants of _dnaB_ will not protect cells from P-lethality.  
Figure 3: _P_-induced plasmid loss. (**A**) Cultures of 594[pclpR-_P_-timm], 594 _clpP_::kan[pclpR-_P_-timm], 594 _grpD55_[pclpR-_P_-timm], and 594[pclpR-_P_\(\Delta\)76-timm] were grown to stationary phase in TB plus 50 \(\mu\)g/ml ampicillin for 48 hr at 25 \({}^{\circ}\)C. Cell aliquots were diluted into fresh TB medium (no ampicillin) as shown in the outline (A) and incubated for about 20 hr in shaking bath between 30 to 42 \({}^{\circ}\)C (refer to Experimental Section _3.8_). (**B), (C**) Plasmid retention by culture cells (described in (**A**)) grown between 30 to 42 \({}^{\circ}\)C.