|
Title Cloning and Expression of Gene Coding For KAC5 from Lactobacillus Reuteri KUB-AC5 by Food Grade Vector System |
|
Type Poster Presentation |
|
Theme Probiotics and Prebiotics: Excellence in Science and Clinical Translation |
|
Topic Development of Probiotic and Prebiotic Foods, Medical Foods, Supplements and Drugs |
|
Main Author Siriphan Sobanbua1 |
|
Presenting Author Siriphan Sobanbua1 |
|
Co-Author Suttipun Keawsompong1 Sunee Nitisinprasert1 |
|
Department / Institution / Country Biotechnology / Specialized Research Unit: Prebiotics and Probiotics for Health, Kasetsart University / Thailand (ไทย)1 |
|
Background and Rationale Background: Lactobacillus reuteri KUB-AC5 isolated from chicken intestine was verified as probitotics which produced antimicrobial peptide (AMP of 4.7 kDa inhibiting various pathogenic bacteria, especially Salmonella serovar Enteritidis S003. Gene coding for AMP was cloned into E. coli by genomic library resulting in a possible open reading frame consisting of 153 and 630 nucleotides encoding 50 and 209 amino acids, respectively. To find out the gene coding for AMP, sub-cloning tecnique was further performed.
|
|
Objectives: Indicates the purpose of the study Objective: This study aimed to localize for the gene encoding AMP by food grade vector system using lactobacilli as host cells. The character of an active AMP was also presented. |
|
Methodology: Describe pertinent experimental procedures Methodology: Gene coding for AMP was sub-cloned and expressed in Lb. plantarum TG02 and Lb. reuteri KUB-AC5 by food grade vector pSIP609. The positive clone of both bacteria was screened by colony PCR method and inhibition activity determination by well diffusion method using S. Enteritidis S003 as an indicator strain. |
|
Results: Summarize the results of the research Results: Subcloning into Lb. plantarum TG02 resulted in an active recombinant clone namely ACLP-C46-F2.1 consisting of 153 nucleotides and exhibiting intracellular inhibition activities of 14.6 mm named KAC5 under induction system of IP-673. It was tolerant at wide pH range of 2-9, high temperature up to 121°C and exhibited activity against both G+ and G- bacteria except lactic acid bacteria. The DNA fragment, I-C46-F2.1, was further subcloned into the wild type Lb reuteri KUB-AC5 resulting in the recombinant clone Lb. reuteri ACLR-C46-F2.1 exerting high activities of 400 AU/ml without IP-673 inducer. The KAC5 would be developed and applied for food and feed safety in the future. |
|
Conclusions: State the main conclusions Conclusion: Overexpression of KAC5 from the recombinant Lb. reuteri ACLR-C46-F2.1 displayed higher inhibition activities for 1.6 folds comparing to the wild type strain. KAC5 characters were wide inhibition spectrum and tolerance to a wide pH range and high temperature. |