Using DNA barcoding to identify the genus Lolium.doc

Using DNA barcoding to identify the genus Lolium Shenmao Wu1, Liping Yin2, Zhirui Deng1, Qin Chen1*, Yining Fu2, Hua jie Xue2 1 Shanghai Key Laboratory of Biotechnology, School of Life Sciences, Shanghai University, No 333, Nanchen road, Shanghai, P.R. China, chenqincc@staff.shu.edu.cn (*corresponding author) 2Technical Center for Animal, Plant and Food Inspection and Quarantine?, Shanghai Entry-Exit Inspection and Quarantine Bureau, Shanghai, P.R. China, yinlp@shciq.gov.cn Abstract. Seeds of the genus Lolium are difficult to identify based on morphology. DNA barcoding has been used to discriminate a variety of agricultural crops. In this study, DNA barcodes for the genus Lolium were investigated. DNA sequences of psbA–trnH, rbcL, atpF–atpH, and the ITS2 region were evaluated for their ability to differentiate Lolium from the related genus Festuca. The greatest divergence existed in ITS2, as confirmed by inter–intraspecific divergence and Kimura 2 parameter analysis. ITS2 could effectively discriminate all species based on a neighbor-joining tree. Thus, the ITS2 region is a candidate for DNA barcoding of Lolium. Keywords: Lolium; DNA barcoding; ITS2; psbA–trnH; rbcl  Introduction Lolium, a genus of Poaceae, consists of six species and one variety: Lolium perenne, Lolium persicum, Lolium remotum, Lolium multiflorum, Lolium rigidum, Lolium temulentum, and Lolium temulentum var. arvense (Raven and Zhang, 2013). Lolium perenne and L. multiflorum are widely used as cool-season forage and have high economic value. However, L. temulentum, when infected by fungus, can produce the toxic alkaloid temulin, which can poison livestock and humans, occasionally resulting in death. Lolium temulentum and its variety are morphologically similar to L. perenne, L. multiflorum, and L. rigidum and cannot be distinguished routinely. In addition, the genus Festuca is closely related to Lolium, and their gene structures indicate they shared a common ancestor about 2.8 million years a