Genome analysis of Lilium tigrinum by chromosome microdissection and molecular cytogenetic techniques

Chromosome microdissection and microcloning are powerful tools for plant genome research. Here we describe the isolation of chromosome #1 derived sequences from L. tigrinum with these techniques and their characterization. Detailed chromosome analysis was performed by FISH and then chromosome #1 was isolated from metaphase chromosomes of L. tigrinum by microbeam dissection. DOPPCR and LA-PCR were used to amplify a DNA of chromosome #1 segments. PCR products from the microdissected chromosome were cloned into a plasmid vector to construct a chromosome #1 specific library and sequenced. BLAST-nr revealed that 28% of the sequences were matched with known genes and transposons, and the rest of 72% did not match with known sequences from NCB1 database of plant taxa. The unknown sequences were putatively divided into five classes and we called them lily unique unknown repeats. FISH confirmation with some clones confirmed that the products from both methods were indeed amplified from the chromosome #1 of L. tigrinum genome. These results provide important information for not only the composition of the Lilium genome but also for detailed sequence information of huge genome sized plants.