OBJECTIVE: To construct a recombinant Lactobacillus acidophilus that expresses high levels of Helicobacter pylori (Hp) adhesin Hp0410. METHODS: The gene fragment encoding Hp0410 was amplified by PCR from the DNA of H. pylori NCTC11639 strain and cloned into the shuttle plasmid pMG36e to construct pMG36e-Hp0410, which was transformed into Lactobacillus acidophilus by electroporation. The target protein was confirmed with SDS-PAGE and silver nitrate staining and analyzed by Western blotting. The stability of the recombinant plasmid was assessed by drawing the growth curve of the recombinant Lactobacillus acidophilus. RESULTS: A 750-bp fragment was inserted into the pMG36e plasmid and transformed into Lactobacillus lactis. The transformed bacterium expressed the target protein with a relative molecular mass of about 34 kD. Western blotting confirmed that the expressed proteins could be recognized by the serum of patients with Hp infection. The recombinant plasmid pMG36e-Hp0410 exhibited good stability in the presence or absence of erythromycin. CONCLUSIONS: The recombinant Lactobacillus acidophilus with high constitutive expression of Hp0410 has been constructed successfully.
OBJECTIVE: To construct a recombinant Lactobacillus acidophilus that expresses high levels of Helicobacter pylori (Hp) adhesin Hp0410. METHODS: The gene fragment encoding Hp0410 was amplified by PCR from the DNA of H. pylori NCTC11639 strain and cloned into the shuttle plasmid pMG36e to construct pMG36e-Hp0410, which was transformed into Lactobacillus acidophilus by electroporation. The target protein was confirmed with SDS-PAGE and silver nitrate staining and analyzed by Western blotting. The stability of the recombinant plasmid was assessed by drawing the growth curve of the recombinant Lactobacillus acidophilus. RESULTS: A 750-bp fragment was inserted into the pMG36e plasmid and transformed into Lactobacillus lactis. The transformed bacterium expressed the target protein with a relative molecular mass of about 34 kD. Western blotting confirmed that the expressed proteins could be recognized by the serum of patients with Hp infection. The recombinant plasmid pMG36e-Hp0410 exhibited good stability in the presence or absence of erythromycin. CONCLUSIONS: The recombinant Lactobacillus acidophilus with high constitutive expression of Hp0410 has been constructed successfully.
Authors: Gayatri Vedantam; Joshua Kochanowsky; Jason Lindsey; Michael Mallozzi; Jennifer Lising Roxas; Chelsea Adamson; Farhan Anwar; Andrew Clark; Rachel Claus-Walker; Asad Mansoor; Rebecca McQuade; Ross Calvin Monasky; Shylaja Ramamurthy; Bryan Roxas; V K Viswanathan Journal: Front Microbiol Date: 2018-09-05 Impact factor: 6.064