Forensic botanical comparison can be hampered by the lack of appropriate DNA databases. While DNA sequence databases for many mitochondrial loci have been established for the identification of animal species, less is known regarding the genomes of plants. We report on the use of the trnL intron and the trnL-trnF intergenic spacer (IGS) in the chloroplast genome and establish a DNA sequence database for plant species identification. The DNA sequences at these two loci from commonly encountered plants, including monocots and dicots, were aligned to establish a DNA database of local plants. The database comprises 373 individual sequences representing 80 families, 206 genera and 269 species. These plant species can be grouped to species level using both sequence and length polymorphisms at these loci. To validate the database for future forensic purposes, we sequenced 20 blind samples and searched the local database and the databases of GenBank and EMBL. Fifteen of these 20 samples used in blind trial testing matched their respective species from our local DNA database but only 6 matched species registered in the GenBank and EMBL databases. The sequences of two species used in the blind trial did not match any sequence registered in any of these databases. Cluster analysis was performed to demonstrate the family and genus distribution of samples. Neighbor-joining trees of the two DNA regions from 70 samples of the local database and 10 of the species used in the blind trials were constructed and clustered to both family and genus. The bootstrap values of the trnL intron were higher than most of those of the trnL-trnF IGS. The sequence database described in this study can be used to identify plant species using DNA sequences of the trnL intron and trnL-trnF IGS of chloroplast genome and illustrates its value in plant species identification.