Screening Of Ramie Germplasms Tolerant To Low Phosphorus And An Improvement Method Of Protein Extraction For Mature Leaves Of Perennial Plants

As an important fiber crop, forage crop and industrial raw material, ramie(Boehmeria L. Gaud) has a pivotal position in Chinese textile industry, animal husbandry and industry. With the adjustment of planting structure, ramie planting area transfers to hillside and barren land, exploring the physiological mechanism of the resistance and stress tolerance in ramie becomes a new research hotspot. Screening aimed material is the basis of studying physiological mechanism of tolerance to low phosphorus in Ramie. In this paper, 138 cultivars were tested by measuring physiological indices which were related to tolerance to low phosphorus, 19 cultivars were tentative screened for low phosphorous tolerant genotypes of ramie, the 19 cultivars were low phosphorus stressed by sand culture, determination of tolerance to low phosphoru was identified by agronomic traits. Based on SSR technology, the genetic diversity of 19 cultivars was analyzed by 39 pairs of primers which were related phosphorus stress.Phosphorus as an important component of many compounds, is a movable element in the process of plant growth which can be able to transport from mature leaves to new leaves, but differential proteomics about mature leaves were rarely reported. To offer technical support for future research related to differential proteomics about phosphorus stress, this study modified the protein extraction method of mature leaves from six common perennial species.The results are as follows:1. In the period of vigorous growth, indicators including relative conductivity, MDA, acid phosphatase activity, total phosphorus were measured in 138 cultivars. Clustering analysis by SPSS 19.0 software showed that, 138 cultivars were grouped into 19 clusters, the number of species in each cluster was 16, 24, 1, 19, 3, 9, 17, 2, 2, 1, 2, 2, 1, 18, 10, 6, 2, 2, 1. Representative species were selected from each cluster, they were “Dianjiangbaima”, “Wangcangwujiaoma”, “Dahuangma”, “Xiaoyelv”, “Huangpigun”, “Puqidayema”, “Xiyelv”, “Dayelv”, “Honghuqingma”, “Hefengzhuma”, “Huazhu 5”, “Xinyuma”, “Xiangzhu 1”, “Zixingma”, “Niuerqing”, “Brazil 8”, “Brazil 3”, “Brazil 2”, “Huangjingpi”.2. Relative tolerance indexes including height, number of green leaves, green leaf area, chlorophyll content, root fresh weight, biomass, fresh weight, root to shoot ratio were measured in 19 cultivars under low phosphorus. Clustering analysis by SPSS 19.0 software showed that, 19 cultivars were grouped into three categories. The analysis result indicates that, “Wangcangwujiaoma”, “Dianjiangbaima”, “Huazhu 5”, “Huangpigun”, “Xiaoyelv”, “Puqidayema”, “Brazil2” were low-phosphorous tolerant genotypes; “Brazil 8”, “Brazil 3”, “Honghuqingma”, “Hefengzhuma”, “Huangjingpi”, “Niuerqing”, “Dahuangma” were low-phosphorus middle resistant genotypes; “Dayelv”, “Zixingma”, “Xiyelv ”, “Xinyuma”, “Xiangzhu 1” were low-phosphorus non-resistant genotypes.3. Genetic diversity analysis of the 19 cultivars by 39 pairs of SSR primers related to phosphorus stress showed that, the 19 cultivars were clustered into four categories, and “Dayelv” was separate for one clade, the genetic relationship with all other species was distant. “Huazhu 5”, “Honghuqingma”, “Hefengzhuma”, “Xiyelv”, “Puqidayelv” were classified into one clade, and the genetic relationship between “Huazhu 5” and “Honghuqingma” was very close. “Brazil 8”, “Brazil 3”, “Brazil 2”, “Xiangzhu 1”, “Zixingma”, “Xinyuma”, “Huangjingpi”, “Wangcangwujiaoma”, “Niuerqing” were classified into one clade, and the genetic relationship between “Brazil 8” and “Brazil 3” was very close. “Xiaoyelv”, “Huangpigun”, “Dianjiangbaima” were classified into one clade, and the genetic relationship between “Xiaoyelv” and “Huangpigun” was very close.4. Based on conventional protein extraction method from young leaves, in this paper, the protein extraction method of mature leaves from six common perennial species was modified. Tris and Triton X-100 were added instead of CHAPS into the lysis buffer, and a 20% TCA-water solution and 100% precooled acetone were added after the protein decomposition for the purification of the protein. The results showed that improved protein extraction methods made two-dimensional electrophoresis patterns more clearly, the number of identified proteins was increased, tailing phenomena was obviously reduced, but the number of acid-soluble proteins was reduced.