1. Graduate University of Advanced Technology, Kerman, Iran 2. Graduate University of Advanced Technology, Kerman, Iran 3. Institute of Biotechnology, Shiraz University, Shiraz, Iran
Abstract
Introduction
Cecropins are a group of antimicrobial peptides (amp) derived from innate immune system of various multicellular organisms. these peptides suppress bacterial infections but have minimal toxic and allergic side effects to the host. thus, antimicrobial peptides may potentially be applied in medicine and agriculture as new and safe antibiotics or bio-control agents. in order to explore pharmaceutical potential and medical importance of antimicrobial peptides, a cost-effective and scalable method to produce large quantities of active peptides is required, and this may be achieved by cloning and expression of recombinant peptides. hairy roots cultivation provides rapid evaluation of peptide activity as well as efficiency and yield of peptide production.
Methods
in the present study, the cecropin a gene was optimized according to codon usage nicotiana tabacum l. and synthesized by generay company. then, constructed vector transferred to the agrobacterium rhizogenes strain atcc 15834 by electroporation and leaf disk techniques respectively. transgenic hairy roots were confirmed with dna and synthesized cdna using specific primers. the total soluble protein (tsp) from the transgenic hairy roots were extracted on stone method and construction was measured using bradford and was detected using anti-his-tag in elisa. the different volume of supernatant (10, 20, 40 and 80 μl) were loaded on 6 mm sterile paper discs.
Results
The antibacterial activity against escherichia coli, aspergillus niger and staphylococcus aureus was evaluated by measuring the diameter of the inhibition zone around the discs (mm).
Conclusion
Cecropin a show inhibitory effect and suitable antimicrobial activity against both gram-positive and gram-negative bacteria.
Keywords
Antimicrobial peptide, cecropin a , nicotiana tabacum
