## 4 Discussion  
We show here that transcription is a major cause of the chromosomal degradation seen in _recA_ cells. The chromosome content defects of cells lacking either Rep [6] or RecA (Figure 1A, compare (iii) with (vii)) share the same primary cause therefore indicating that both Rep and RecA reduce the impact of gene expression on genome duplication. The synergistic increase in chromosome content defects in _rep recA_ cells indicate that these enzymes provide alternative means of mitigating the impact of transcription on DNA replication (Figure 1A (iv)). Furthermore, the significant suppression of DNA degradation in _recA_ cells by an RNA polymerase mutation supports the view that protein-DNA complexes are the primary causes of replication defects in cells not exposed to elevated DNA damage [6]. However, the time taken to duplicate a chromosome is not extended in the absence of either RecA or RecB, in contrast to cells lacking Rep (Figure 3). Thus the maintenance of rapid chromosome duplication has a greater dependency on Rep as opposed to RecA or RecBCD. RecA and RecBCD do, though, have the ability to sustain chromosome duplication in _xrep_\(\Delta\)_uvrD_ double mutant cells when transcriptional barriers to replication are reduced (Figures 4 and 7). Both RecA and RecBCD are needed for this underpinning, demonstrating that maintenance of chromosome duplication by recombination enzymes is most efficient when RecBCD catalyses loading of RecA at dsDNA ends rather than large-scale RecBCD-dependent degradation of such ends (Figure 7). These data are apparently contradictory. RecA has little impact on chromosome duplication times and, although the time resolution of the measurements in Figure 3 are relatively low, they still imply infrequent engagement of RecA in genome duplication during a single cell cycle. In contrast, the absence of RecA results in frequent chromosome degradation [55] and a reduction in viability [85]. In considering this apparent contradiction, there are many factors that potentially affect the time needed to copy a chromosome (Figure 8). The inherent speed and processivity of the replisome is an important determinant of chromosome duplication time but the pausing behaviour of replisomes, and what happens to these paused forks, will also impact on duplication times (Figure 8 (i-v)). Figure 6: RecB is essential in _rep uvrD propB*35_ cells on rich medium. (**A–E**) Loss of PAM375 (pRC7_rrcB_) from the strains indicated was monitored on LB Xgal IPTG. Figure 7: The requirement for RecBCD and RecA is not alleviated by mutation of _recF_. (**A–D**) Loss of PAM406 (pRC7_rrcA_, _recB_) from the strains indicated was monitored on LB Xgal IPTG. Accessory helicases reduce the frequency and/or duration of replisome pauses at nucleoprotein complexes and increase the probability of paused replisomes restarting replication as opposed to losing function [6; 22; 27]. The extended chromosome duplication time in _exp_ mutant cells [29; 31] indicates that one or more of these pausing parameters are critical in determining the speed of chromosome duplication. The more than twofold increase in chromosome duplication times in _exp_ mutant cells probably also underrepresents the significance of replisome pausing behaviour on these timings, given the ability of UvrD to compensate partially for the absence of Rep [22; 30]. Regardless of the cause of replisome pausing, there is no evidence that recombination enzymes impact directly on paused replisomes but they can act after a replisome has lost function to promote replication restart [7]. Loss of enzymes such as RecA or RecBCD might therefore impact on the duration of any fork repair process since loss of an enzyme normally involved in fork repair might lead to extension of repair times due to less efficient alternative pathways that are not normally operative in wild type cells (Figure 8 (vi)). A related consideration is the position of replication re-initiation with respect to the position of the blocked initial replisome (Figure 8 (vii)).