Temporins,antimicrobialpeptidesfromtheEuropeanred.pptVIP

Temporins, antimicrobial peptides from the European red frog Rana temporaria 05基地班 魏潇 冯秀云 程超 靳利蕊 张蓉 卢会文 Introduction Skin secretions of frogs contain many different types of antibacterial peptides. In particular, a variety of such peptides has been isolated from several Rana species. There are 4 different groups: brevinin 1 Family, brevinin 2 Family, 37-residue peptides, a group that consists of esculentin 1--has the highest antibacterial activity. The signal sequences of precursors of antimicrobial peptides from Rana species were very similar to those with different biological activities from Phyllornedusa species. Search for the precursors of new peptides in the skin of Rana temporuria, whose signal peptide is similar to signal peptide of esculentin 1. Methods And Result Isolation of RNA and cDNA screen Northern blot analysis Collection and purification of skin secretions and Structural analysis Antimicrobial assay Isolation of RNA and cDNA screen Isolation of poly(A)-rich RNA by affinity chromatography Preparation of the cDNA library Preparation of probe: a 240-bp fragment obtained by digestion of the esculentin 1 cDNA with HindIII, then labelled by random priming. Screening Positive clones were selected and analysed by cleavage with restriction enzymes and nucleotide sequencing. Result Six positive clones were detected. The sequences of the inserts present in clones Rt-5, Rt-6, and Rt-17 are shown in Fig. 1. Northern blot analysis Poly(A)-rich RNA (5 μg) was fractionated by electrophoresis Blotted directly onto Nytran sheets Probe: The insert of clone Rt-17 labelled by random priming. Hybridization Result the probe derived from clone Rt-17 recognized an abundant message, detected as a single, rather broad band in the range 400-500 nucleotides. Under the same conditions, no signal could be obtained from the skin of other amphibian species. Collection and purification of skin secretions and structural analysis Three specimens of R. temporaria (30-35 g each); Main