A simple method to provide a shuttling plasmid for delivery to other host ascertained by prolonged stability of extracellular plasmid DNA released from Escherichia coli K12 endA mutant, deficient in major endonuclease.
Escherichia coli lyses by lambda phage propagation. Circular plasmid DNA was present during E. coli lysis as an extracellular plasmid DNA (excpDNA) that was stable enough to transform coexisting competent Bacillus subtilis cells. Detailed investigations unveiled that excpDNA is transient in both quality and quantity, with stability lasting no more than several hours. A survey using E. coli lambda lysogens with various genetic backgrounds demonstrated that the loss of Endonuclease I (ΔendA::kan) conferred extraordinary stability upon excpDNA for as long as 48 h. Studies on endA mutants suggested that excpDNA remained localized in cell debris, in contrast to E. coli genome DNA, which diffused into medium at an early point in lysis. Lambda lysogens constructed on endA recA mutants are presented for potential pipelines in delivery to other competent proficient microbes.
Institute for Advanced Biosciences, Keio University, Nipponkoku, Tsuruoka, Yamagata 997-0017, Japan. mita2001@sfc.keio.ac.jp
Mitsuhiro
M
Itaya
Yoko
Y
Kawata
Mitsuru
M
Sato
Masaru
M
Tomita
Kenji
K
Nakahigashi
D001412:Bacillus subtilis / Q000235:genetics*; D010582:Bacteriophage lambda / Q000235:genetics / Q000502:physiology; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000235:genetics* / Q000378:metabolism; D048168:Escherichia coli K12 / Q000201:enzymology / Q000235:genetics* / Q000821:virology; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D055786:Gene Knockout Techniques; D008242:Lysogeny / Q000235:genetics; D009154:Mutation; D010957:Plasmids / Q000235:genetics* / Q000378:metabolism; D011938:Rec A Recombinases / Q000235:genetics; D014169:Transformation, Bacterial; D014779:Virus Replication
D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D011938:Rec A Recombinases; D003850:Deoxyribonuclease I