[Development of DUS test guidelines for new varieties of Gynostemma]

To meet the urgent needs for variety breeding and variety-right protection in the genus Gynostemma, this study developed the guidelines for the testing of distinctness, uniformity, and stability(DUS) for new varieties of this genus in accordance with GB/T 19557.1 and relevant international standards. A total of 58 Gynostemma germplasm resources were collected from nine provinces, covering two major applied species(G. longipes and G. pentaphyllum) and 10 other species, including G. burmanicum and G. yixingense. Field trials were conducted in Zhenping county, Shaanxi province, where all germplasms were uniformly planted using asexual propagation. Through observations across the entire growth cycle, trait expression at each developmental stage were systematically recorded, and 42 traits were screened, encompassing rhizomes, stems, leaves, flowers, fruits, seeds, and other organs. These traits included 5 qualitative traits, 31 quantitative traits, and 6 pseudo-qualitative traits, as well as 29 visual traits, 11 measurable traits, and 2 traits evaluated using both methods. Variation analysis of 12 measurable traits showed coefficients of variation ranging from 8.42% to 53.83%, with the highest value observed for rhizome dry weight(53.83%). Most traits exhibited significant differences among varieties. Correlation analysis indicated that only 31 of the 66 trait pairs exhibited significant or highly significant correlations, and correlations among most traits were weak, thus avoiding redundancy in discriminatory power. Measurable traits were graded using the median-average standard deviation method combined with the least significant difference(LSD) method. The contents of gypenoside A, gypenoside XLIX, and total gypenosides were incorporated into the testing system, and nine standard varieties were initially selected to correct for environmental effects. This study established a systematic trait evaluation system for the genus Gynostemma, helping to address challenges arising from interspecific morphological similarity and difficulties in identification. It provides scientific and technological support for the identification, breeding, and protection of new Gynostemma varieties, and also serves as a reference for the development of DUS testing guidelines for medicinal plants.