DNA sequencing and comparative sequence analysis reveal that the Escherichia coli genomic DNA may replace the target DNA during molecular cloning: evidence for the erroneous assembly of E. coli DNA into database sequences

Comparative Biochemistry and Physiology. Part B, Biochemistry & Molecular Biology
P K Jha, S Sarkar

Abstract

DNA sequencing and similarity search of databases provide experimental evidence that portions of the host Escherichia coli genome may get ligated into the cloning vector, resulting in clones containing nontargeted inserts. Several lines of evidence suggest that this non-targeted ligation, as observed by us while subcloning troponin I cDNA, is presumably due to a recombination-mediated mechanism by which host DNA replaces the target DNA in the cloning vector. The E. coli genome mapping to 64-65 min and 92.8-00.1 min, the latter containing insertion sequences, appears to be the hotspot regions involved in this process. We examined the possibility that some sequences reported in the databases may also contain genomic sequences of E. coli. A search of current databases revealed that a rat hepatic glutathione transporter cDNA contains a 2.2-kb-long portion of the E. coli genome that has been wrongly assembled into its 5' untranslated and coding regions. In addition, about 30 sequences in databases, including a Yersinia pestis toxin gene, showed relatively high sequence identity with those portions of the E. coli genome that were present in the nonauthentic clones.

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Citations

Jul 9, 2004·BMC Evolutionary Biology·I King JordanEugene V Koonin
Sep 17, 2003·BMC Genomics·Olga Zhaxybayeva, J Peter Gogarten
Oct 17, 2012·Parasitology·Mercedes AlonsoMontserrat Espiñeira
Jun 5, 2019·Proceedings of the National Academy of Sciences of the United States of America·Li LiuTian'en Zhang
Jun 30, 2018·The Journal of Microbiology·Leonid N TenHee-Young Jung
Aug 10, 2021·PLoS Computational Biology·Nikolas DovrolisIoannis Karakasiliotis

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