Taxonomic Study On The Genus Sanicula(Apiaceae)from China

Sanicula L.represents a monophyletic group in the family Apiaceae with a worldwide distribution of approximately 40 species,17 of which occur mainly in southwestern China.The interspecific relationships of Chinese Sanicula have heretofore been difficult to resolve.In conjunction with chloroplast genome studies and DNA sequencing,fruit and pollen morphology were examined to explore the interspecific relationships of Chinese Sanicula and further our phylogenetic understanding of the Apiaceae family.The availability of more than 7,000 collections of Sanicula species from North America and Asia laid the foundation for this research and provided materials for examination.Fifteen Sanicula species were included in the palynological study.Pollen grains from each species were examined by Light Microscopy(LM)and Scanning Electron Microscopy(SEM).These pollen grains vary tremendously in shape.Their basic structure was trigonal,tricolporate,pleurotreme,with a2-layered exine.The statification of exine was clear.The pollen was divided into two clearly distinguishable types: rectangular or equatorial constricted.These results support Sanicula as a recently evolved group in Apiaceae.Within Sanicula,taxa having larger pollen grains and a straight equator represent more recently evolved taxa and those possessing smaller pollen grains with a contracted equator represent more primitive taxa.Our results indicate that Sanicula chinensis and S.elata are most primitive within the genus.Fruit micromorphology of the 15 Sanicula species studied was also informative.The size of the fruits were found to be 1.5-3.2 mm × 1.5-2.8 mm,with significant differences among species such as S.gregaria Bicknell(1.5 ×1.5mm)and S.marilandica L.(3.2 × 2.8mm).Three micro-morphology characters of the spine were examined: spine wax type,wax density,and wax flatness.Differences among taxa for these characters proved to be slight and consequently insignificant.Seven micromorphological characters of theridges of the surface protrusions and the fruit pedicles were also studied.These characters included protuberant type,protuberant level,and protuberant cell shape and wax excretion characters which included wax type,wax density,wax flatness,and relative length of wax.These micromorphological characters among taxa proved to differ significantly and were included in a newly established identification key to Sanicula taxa.In order to reconstruct the phylogeny of Sanicula,the nuclear ribosomal DNA ITS gene was sequenced in 11 Sanicula taxa as well as from representatives of Eryngium and Hacquetia in the Saniculoideae.Results support previous work demonstrating that the genus Sanicula is a monophyletic group(BS=100%)and sister to Eryngium(BS=82%).Sanicula rubriflora and Hacquetia epipactis are supported as monophyletic(BS=80%).Therefore,H.epipactis has been transferred to Sanicula epipactis(Scop.)E.H.Krause.Reported here for the first time are the chloroplast genome sequences of Sanicula chinensis and S.orthacantha.Genome sizes are 154,650 and 157,163 bp,respectively,and each has a typical quadripartite structure with the large(LSC)and small(SSC)single-copy regions separated by a pair of inverted repeats(IRs).Their gene content and organization are considerably similar to previously reported members of Apiaceae.Our results show that the following regions in the Sanicula chloroplast genome are highly divergent: matK,rps15,ycf2,rps13,ndhA,petB,petN,psbL,trnS-trnG.Using the Maximum Likelihood(ML)method to construct the phylogenetic trees,our results support previous research and show that Sanicula is closely related to other Apioideae(BS=100%)and sister group to the Hydrocotyloideae(BS= 100%).