Selection of highly variable chloroplast DNA sequences for genetic relationship study of tobacco cultivars in Thailand

Abstract

In Thailand, imported and local tobacco cultivars have different regulations in tariff collection. However, there are major legal and technical problems on how to distinguish between the two groups of tobacco cultivars. In this study, highly-variable noncoding regions of chloroplast DNA was ntroduced to study genetic relatonships among totally 51 local and imported tobacco cultvars (Virginia, Turkish, and Burley cultivar groups) grown in Thailand. The amplification and sequencing results revealed that six out of nine regions (rp/32-trnL, ndhC-trnV, ndhF-rpl32, psbD-trnT, psbJ-petA and atpl-atpH) showed polymorphic sequence characteristcs, both in sequence engths and the amounts of base usbstitutions. Flmost all of Virginia cultivars were different in nucleotide sequence from the other cultivars. rpl32-trnL region could also distinguish seven local cultivars and one roll-your-own tobacco product with a large 66 bp insertion. The sequence polymorphism of rpl32-trnL region was in the highest vaue with 10.47% variability and 75 potentially informative characters. A multiplex PCR technique was successfully developed to distinguish Virginia cultivars and the seven special local cultivars from the others. This marker can be used as a solution for the "local" and "imported" cultivar problem.