Knocking out Analysis of the CpxP gene using Crispr/Cas9 in Escherichia
Based on the analysis of CpxP genes among Escherichia coli strains, CpxP
gene-targeting short guide RNA (sgRNA) was designed and inserted into
the pGL3-MGP-RNA. The donor sequences (MG-HR) for homologous repair were
designed and cloned by PCR. MG-HR and pGL3-MGP-RNA were transformed into
E. coli MG1655 (pCas9). The CpxP gene expression cassette was amplified
by PCR and subcloned into pBBR1MCS-2. Then the pBBR-CpxP was
independently transformed into E. coli MG1655. The results of motility
experiment suggest that CpxP gene had a significant effect on the
movement ability of E. coli strain. The CpxP protein had a significant
inhibition of bacterial activity. The lastest 81 CpxP proteins sequences
were selected and analyzed by multi-sequence alignment and molecular
cluster. The CpxP proteins were roughly divided into three categories.
Our results suggest that the CpxP protein was involved in bacterial
motility, infection and pathogenicity.