Biology subjects

Sanchez Olmos, M. d. C.

Publications and source records attributed to Sanchez Olmos, M. d. C..

2 recordsLinked to original sources

bioRxiv2023

DNA replication is not a limiting factor for the rapid growth of Vibrio natriegens

DNA replication prior to cell division is essential for the proliferation of all cells. Bacterial chromosomes are replicated bidirectionally from a single origin of replication, with replication proceeding at about 1000 bp per second. For the best-studied model organism, Escherichia coli, this translates into a replication time of about 40 min for its 4.6 Mb chromosome. Nevertheless, E. coli can propagate by overlapping replication cycles with a maximum short doubling time of 20 min. The fastest growing bacterium known today, Vibrio natriegens, is able to replicate with a generation time of less than 10 min. It has a bipartite genome with chromosome sizes of 3.2 and 1.9 Mb. Is simultaneous replication from two origins a prerequisite for its rapid growth? We fused the two chromosomes of V. natriegens to create a strain carrying a 5.2 Mb chromosome with a single origin of replication. Compared to the wild-type, this strain showed little deviation in growth rate. This suggests that the split genome is not a prerequisite for rapid growth, and that DNA replication is not an important growth rate-limiting factor.

microbiology↗

bioRxiv2022

L-SCRaMbLE creates large-scale genome rearrangements in synthetic Sc2.0 chromosomes

Optimization of the metabolic flux through heterologous pathways to improve bioproduction or utilization of alternative substrates requires both fine-tuning of non-native gene expression levels and improvement of the host genome. The SCRaMbLE system incorporated into synthetic Sc2.0 yeast strains enables a rapid approach to rearrange the genome of Saccharomyces cerevisiae in order to create optimized chassis. Here, we show that the light-inducible Cre recombinase L-SCRaMbLE can efficiently generate diverse recombination events when applied to Sc2.0 strains containing a linear or circular synthetic chromosome III. We present an efficient and straightforward workflow for the identification of complex rearranged synthetic chromosomes from SCRaMbLEd isolates without selection pressure. The screening method is based on novel genotyping primers, the loxPsym tags, which indicate not only deletions but also inversions and translocations. Long-read Nanopore sequencing is used to decode the selected genotypes and shows in conjunction with flow cytometry that large-scale karyotype alterations can be a consequence of SCRaMbLE.

synthetic biology↗