Identification of PCR-RFLP Haplotypes for assessing genetic variation in the Green Oak Leaf Roller Tortrix viridana L. (Lepidoptera, Tortricidae)

PCR-RFLPs were performed to assess intraspecific variation in the green oak leaf roller, Tortrix viridana. The cytochrome oxidase I and II genes were amplified with universal and self designed primers, respectively, resulting in three PCR-fragments of 802 bp, 729 bp and 680 bp. 29 restrictions endonucleases were tested for variation in these PCR-patterns. Seven of these enzymes were chosen for further research. We found 13 haplotypes in four populations across a total of 436 individuals. In addition all haplotypes were sequenced. More single nucleotide substitutions were detected in the sequences, particularly in the middle of the cytochrome oxidase I gene, missed by the used restriction enzymes. For these markers intraspecific variation in T. viridana is high compared to other insect species. Furthermore we found differences in frequency of haplotypes among the investigated populations which induce that the markers developed so far are suitable for population genetic studies in T. viridana.