「Biophysics and Physicobiology」に Atsuko Uenoyama, Hana Kiyama, Mone Mimura, Makoto Miyata による "Rapid in vitro method to assemble and transfer DNA fragments into the JCVI-syn3B minimal synthetic bacterial genome through Cre/loxP system" をJ-STAGEの早期公開版として掲載
2024年11月07日 学会誌
日本生物物理学会欧文誌[Biophysics and Physicobiology]に以下の論文が早期公開されました。
Atsuko Uenoyama, Hana Kiyama, Mone Mimura, Makoto Miyata
"Rapid in vitro method to assemble and transfer DNA fragments into the JCVI-syn3B minimal synthetic bacterial genome through Cre/loxP system"
URL:https://doi.org/10.2142/biophysico.bppb-v21.0024
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- Abstract
- JCVI-syn3B (syn3B), a minimal synthetic bacterium that only possesses essential genes, facilitates the examination of heterogeneous gene functions in minimal life. Conventionally, Escherichia coli is used to construct DNA fragments for gene transfer into the syn3B genome through Cre/loxP system. However, the construction process is challenging and time-consuming due to various issues, including the inhibition of E. coli growth and unexpected recombination, especially with AT-rich DNA sequences such as those found in Mycoplasma genes. Therefore, in this study, we aimed to develop a new transformation method to overcome these issues. We assembled the vector and target DNA fragments using an in vitro homologous recombination system and subsequently transferred the products into the syn3B genome. We obtained approximately 103~104 recombinant colonies per milliliter of the original culture in eight days, which is four days shorter than the conventional period, without any recombination issues, even for AT-rich DNA. This method may be applicable to other gene manipulation systems based on Cre/loxP system.
URL:
https://doi.org/10.2142/biophysico.bppb-v21.0024