Molecular Identification of Fritillaria taipaiensis and Its Relatives Based on Indel Marker
OBJECTIVE To construct molecularidentification of Fritillaria taipaiensis and its relatives based on Indel marker.METHODS One hundred and three chloroplast genomes of 13 Fritillaria species were downloaded from GenBank and were subse-quently used to screen Indel marker based on sequence alignment.Then,in accordance with the conserved upstream and downstream of Indel marker,a pair of specific primers was designed accordingly,by which a method including routine PCR and electrophoresis was constructed.Furthermore,the specificity,sensitivity,and optimal reaction temperature of the method were assessed.RESULTS One deletion of 137 bp was found in the accD-psaI spacer region of F.taipaiensis chloroplast genome according to the multiple sequence alignments,which was defined as the Indel marker for F.taipaiensis.The constructed method exhibited high specificity due to the fact that only F.taipaiensis exhibited a band of 302 bp after routine PCR and electrophoresis.The limit of detection for F.taipaiensis DNA template was 0.239 ng·μL-1,and the optimal melting temperature(Tm)value was 58 ℃.CONCLUSION This accurate,quick and cost-affordable method provides not only identification of F.taipaiensis and its related species,but also technical reference for identification of other traditional medicines.
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