Naturally transformable bacteria are able to take up DNA to acquire new genetic traits in the environment. To be naturally transformed, bacteria need to establish a physiological state, called natural competence, in which DNA uptake and processing genes are expressed. DNA uptake proteins assemble a complex to pull exogenous DNA into the cytoplasm where it can recombine with the genome DNA or establish as a plasmid. In general, DNA uptake of bacteria could be divided into two stages: DNA is transported from the milieu to the periplasm at the first stage (for Gram-negative bacteria) and is translocated across the inner membrane at the second stage. Our work and other studies revealed new plasmid DNA transformation modes in Escherichia coli. Here, we first reviewed recent advances in the molecular mechanism of natural transformation and then described the distinctive plasmid transformation mode in E. coli.