Genetic and chemical components analysis of Papaver setigerum naturalized in Korea.

Of the 110 species of genus Papaver, only Papaver somniferum and P. setigerum are controlled poppies in Korea. All poppy samples share similar morphology therefore it is important to check if they contain controlled substances such as morphine and codeine for forensic purpose. Since the alkaloid content of Papaver plants varies according to their growing stage, chemical components analysis alone is not enough to identify exact species. In 2010, hundreds of poppy plants suspected to be P. somniferum were found in Jeju Island, South Korea. They had a slightly different but overall similar appearance to P. somniferum. Using GC-MS analysis, codeine, rhoeadine, papaverine, protopine, noscapine, setigeridine and trace amounts of morphine were detected in these samples. Although their chemical components were different from what has been described in literatures for P. setigerum, they could be assumed to be P. setigerum based on their morphological features and GC-MS results. Also, chromosome numbers using their seeds showed 2n=44 and the numbers were in accordance with those of P. setigerum. Nucleotide substitution or insertion/deletion of ITS (internal transcribed spacer), 18S rRNA (ribosomal RNA), rbcL (large subunit of ribulose 1,5-bisphosphate carboxylase), trnL-trnF IGS (intergenic spacer), trnL intron and psbA-trnH were assessed as universal genetic markers for P. setigerum. Also, genetic analysis using six target genes involved in the biosynthesis of benzylisoquinoline alkaloids, including TYDC (tyrosine/dopa decarboxylase), SAT (salutaridinol-7-O-acetyltransferase), BBE (berberine bridge enzyme), COR (codeinone reductase), CYP80B1 ((S)-N-methylcoclaurine 3'-hydroxylase) and NCS (norcoclaurine synthase) were tested as Papaver-specific genetic markers by the existence of their PCR products. From the results, the sequences of the 6 universal genetic markers and 6 Papaver-specific genetic markers for P. setigerum were identified and then Genbank accession numbers of them were registered in NCBI. Also, the trnL intron and psbA-trnH nucleic acid sequences of the 7 Papaver species were identified and registered.

Exploiting expressed sequence tag databases for the development and characterization of gene-derived simple sequence repeat markers in the opium poppy (Papaver somniferum L.) for forensic applications.

Simple sequence repeat (SSR) markers in the opium poppy (Papaver somniferum L.) were identified from an expressed sequence tag (EST) database comprised of 20,340 sequences. In total, 2780 SSR-containing sequences were identified. The most frequent microsatellite had an AT/TA motif (37%). Twenty-two opium poppy EST-SSR markers were presently developed and polymorphisms of six markers (psom 2, 4, 12, 13, 17, and 22) were utilized in 135 individuals under narcotic control investigation. An average of three alleles per locus (range: 2-5 alleles) with a mean heterozygosity of 0.167 was detected. Six loci identified 29 unique profiles in 135 individuals. The EST-SSR markers exhibited small degrees of genetic differentiation (fixation index = 0.727, p < 0.001). Other variable markers will be needed to facilitate the forensic identification of the opium poppy for future cases. To determine the potential for cross-species amplification, six markers were tested in five Papaver genera species and two Eschscholzia genera. The psom 4 and psom 17 primer pair was transferable. This is the first study to report SSR markers of the opium poppy.