The chloroplast trnS-psbC gene regions from total genomic DNA of 119 accessions from seven small millet
species were amplified by polymerase chain reaction (PCR) and digested with eight restriction enzymes individually as
well as in combinations of two enzymes to generate restriction fragment length polymorphism (PCR–RFLP). PCR–
RFLP with individual enzymes revealed polymorphism between only some species. However, all the species could be
distinguished by using a combination of two enzymes, specifically HaeIII and MspI. PCR–RFLP of 11 to 20 accessions
with the same enzyme combination showed no intraspecific variation, which established that the differential banding
patterns were species specific. In contrast, the same enzyme combination was not useful for differentiating different
species of the genera Cajanus, Rhyncosia, Abies, Rhizophora, Ceriops, and Bruguiera, and it also revealed intraspecies
variation in three species of Abies. The present study indicated that digestion of trnS-psbC with two four-base recognizing
enzymes reveals more variation than with either enzyme alone and that it may be a method of choice for species
identification in some genera.

http://www.nrcresearchpress.com/toc/gen/44/3