Studies On Molecular Cytogenetics Of Nelumbo

Nelumbo is an ancient genus, which is considered to be one of the earliest genuses in angiosperm. In classical taxonomy, lotus belongs to Nelumbo, Nyphaeaceae, Ranales. It has two species, i.e. Nelumbo nucifera Gaertn and Nelumbo lutea (wild) per. But there are many discussions on its taxonomic position. Lotus originated in China, and it is beautiful water ornamental and economic plant. It has been widely planted and used. But researches on molecular cytogenetics of lotus were limited. In this study, we studied the molecular karyotype on Nelumbo nucifera and Nelumbo lutea. The single copy genes bzl and bz2 were also physically mapped onto the chromosomes of Nelumbo nucifera. The main results are as follows:1. Prometaphase chromosomes of Nelumbo nucifera Gaertn ( 2n=16 ) were quantitatively analyzed based on their DAPI staining patterns. The DAPI staining showed prominent uneven distribution along the chromosomes. Based on the DAPI staining patterns (from short arm to long arm), with chromosomal relative length and arm ratio as supplementary parameters, the quantitative chromosome map was constructed for the first time in Nelumbo nucifera Gaertn. by the MetaMorph software. The quantitative chromosome map was the intuitional mode of actual premetaphase chromosomes. It could not only identify every chromosome in genome but also analyze fractional area of heterochromatin in chromosomes. The quantitative chromosome map is therefore considered to be a new method for cytological studies of lotus.2. Detailed karyotypes of Nelumbo nucifera gaertn and Nelumbo lutea (wild) per. were compared on the basis of genome size, chromosome arm length, G-like bands, Ag-NOR staining, CMA bands, CPD bands, FISH detection of 45S rDNA sequence and telomere sequence and genomic in situ hybridization (GISH). The features or recognition markers of each chromosome in these two species were identified. The differences we observed between the chromosomes of the two species seem to be within the variation ranges of a species. So according to the results, we consider Nelumbo nucifera and Nelumbo lutea asdifferent subspecies within the same one species instead of different species.3. Both bzl and bz2 are anthocyanin biosynthetic genes, and related to development of aleuronic layers of maize seeds, bzl encodes UDP glucose flavonoid glucosyl-transferase (UFGT), an anthocyanin biosynthetic enzyme in the aleuronic layer of the seed, which modifies purple aleurone and plant color to pale or reddish brown, anthers yellow-fluorescene. bz2 is another anthocyanin biosynthetic gene, its potential function are reflavonoid acylation, glycosylation transport or deposition. Using maize bzl and bz2 gene inserted into pUC19 labeled by the biotin as the probes respectively, bzl and bz2 genes have been assigned to metaphase chromosome by FISH. The red hybridization signals indicated the bzl gene and bz2 gene on homologous chromosome 2 and chromosome 4, respectively. The percent distances from the centromere are 79% and 67%, respectively. The current studies would advance the addition of genetic markers and the construction of genetic map in lotus.In conclusion, innovative results of this study are those that the differences of N. nelumbo and N. lutea has been identified by new developed molecular cytogenetic technique, the evolutional relationship between the two species of the genus has been analyzed at the level of molecular banding and new information about chromosomal morphology and structure has been offered in tested species.