Acidic antibacterial peptides and gene as well as its application in pharmacy
An antimicrobial peptide and acidic technology, applied in the field of biomedicine, can solve problems such as minor side effects, achieve the effect of inhibiting bacterial growth, simple structure, and convenient artificial synthesis
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0044] 1. Acidic antimicrobial peptide gene cloning:
[0045] 1), extraction of total RNA from the skin of Bombina grandis: the living Bombina grandis was cleaned with water, put into liquid nitrogen for quick freezing for 4 hours, the skin tissue was taken, weighed, 300 mg of skin tissue was taken, and 10 ml of total RNA extraction buffer was added ( Trizol solution, product of GIBCOBRL, USA), homogenate in a 20ml glass homogenizer for 30 minutes, add an equal volume of phenol / chloroform solution, mix vigorously, place at room temperature for 10 minutes, centrifuge at 12000rpm for 10 minutes at 4°C, and discard the precipitate , add an equal volume of isopropanol to the supernatant, place it at room temperature for 10 minutes, centrifuge at 12000 rpm for 10 minutes at 4°C, wash the precipitate once with 75% ethanol, and dry it in the air. The sediment at the bottom of the tube is the total RNA of Bombina maxima skin.
[0046] 2) Purification of mRNA from the skin of Bombina m...
Embodiment 2
[0052] 1. Preparation of Acidic Antimicrobial Peptides:
[0053] The preparation method of the acidic antimicrobial peptide: the amino acid sequence of the acidic antimicrobial peptide is deduced according to the gene encoding the bombesin antimicrobial peptide, and the whole sequence is synthesized by an automatic polypeptide synthesizer. Reversed phase C by HPLC 18 Desalted and purified by column chromatography.
[0054] 2. The molecular weight was determined by Fast atom bombardment mass spectrometry (FAB-MS), using glycerol: m-nitrobenzyl alcohol: dimethyl sulfoxide (1:1:1, V:V:V, volume ratio) as substrate, Cs + As the bombardment particles, the current is 1μA and the emission voltage is 25Kv.
[0055] 3. The purity of the purified acidic antimicrobial peptide was identified by high performance liquid chromatography (HPLC), the molecular weight was determined by fast atom bombardment mass spectrometry, the isoelectric point was determined by isoelectric focusing electr...
Embodiment 3
[0058] The role of acidic antimicrobial peptides in inhibiting bacterial growth:
[0059] The antibacterial activity was detected by the cup and saucer method, and the medium was ordinary agar medium. Inject 20ml of heated and melted medium into the plate as the bottom layer, spread it evenly in the bottom of the plate, after solidification, take another appropriate amount of medium and heat and melt, add 5ml of bacterial suspension to each plate, shake well, Spread it evenly on the bottom layer as a bacterial layer. After cooling, put 6 sterilized stainless steel cups evenly in the plate at equal distances. Add 0.1ml of the test compound solution with a concentration of 0.1-0.3mg / ml to the first steel cup, add the sample solution to the other steel cups by doubling dilution method, incubate at 37°C, and observe the size of the inhibition zone. The bacteriostatic zone above 10mm was regarded as the minimum inhibitory concentration (minimal inhibitory concentration, MIC). Th...
PUM
Login to View More Abstract
Description
Claims
Application Information
Login to View More