SBIR-STTR Award

There is a need for locally-adapted native plants which are tolerant of very low soil pH and elevated heavy metal concentrations to re-vegetate acid mine land in the Rocky Mountain west. Large numbers of acid and heavy metal tolerant (AHMT) plants are necessary because of the immense area of acid mine land that needs to be reclaimed. In Montana alone there are about 6000 abandoned hard rock mines and milling sites. Mining wastes have contaminated over 3,000 miles of rivers in Western states. AHMT plants will be of value even on amended or cover-soiled sites as plant roots often extend into un-amended soil. Researchers have experimented with seed collected from such sites as a way to produce AHMT locally adapted native plants. However, many desirable native plants are difficult to propagate by seed or by vegetative production (e.g. cuttings). Plant tissue culture is a way to overcome these problems, but few of the most desired plants and no AHMT plants have ever been cultured before. The purpose of this research is to produce AHMT plants using plant tissue culture. SMK Plants will collect plant material for AHM sites, develop tissue culture propagation methods for these plants, and seek to attain commercial production rates to provide cost competitive AHMT plants for mine land reclamation. Revegetation of mined land with these plants will benefit society by reducing wind and water erosion which in turn will reduce toxic materials in the air we breathe and the water we drink. OBJECTIVES: The objective of this research is determine the feasibility of using plant tissue culture to produce large numbers of site-adapted, native plant species useful for acid/heavy metal (AHM) contaminated mine sites. First, SMK Plants will select a range of plant species determined to be valuable for revegetating a representative AHM contaminated mine site in Montana. Second, we will secure suitable AHM contaminated site(s) that contain the plant species for collection. Third, we will utilize the plant tissue culture, or micro-propagation, process to determine if plant tissue culture can be used to propagate nursery quality plants of these collections in quantities for commercial revegetation operations. APPROACH: With the aid of highly respected consultants in the reclamation field, SMK Plants will select the species to be studied and secure the AHM site in Montana for collection. The plant tissue culture process will involve protocols developed and/or utilized by SMK Plants for propagating field collected native species. It will proceed as follows: (1) Collect portions of plants(explants) of the selected species. Identify each plant or population with GPS coordinates and physical markers; (2) To ensure that the plants from which the explants are taken are in fact AHMT, take soil samples from the root zone of the plants from which the explants were taken and have them analyzed; (3) Disinfest and establish each of the selected plants in clean (i.e. no fungal or microbial contaminants) culture; (4)Establish shoot cultures and determine media and growing conditions to achieve a commercial multiplication rate for each of the selected plants, i.e. 4 to 6 X in 4 to 6 weeks; (5) Test in vitro rooting method to achieve rooting within 6 weeks after transferring to rooting media. Finally, produce 1000 tissue cultured plants for each species and determine the unit cost of production

SMK Plants LLC will use plant tissue culture to quickly produce large numbers of acid-soil heavy-metal tolerant (AHMT) locally-adapted native plants suitable for revegetating land contaminated by hard rock mining wastes. Large numbers of AHMT plants are in demand because of the immense area of degraded mine land needing to be reclaimed. It will take billions of dollars to reclaim these lands. We proved in SBIR Phase I the principle that we can find, collect, and tissue culture native plants which grow directly on acidic heavy-metal (AHM)contaminated soil. These plants multiply and root similarly to commercial ornamental plant species with similar production costs, making them economical for reclamation. The goal for our SBIR Phase II project is to tissue culture locally adapted native plant species growing on AHM soils, then successfully grow and over-winter them under commercial greenhouse conditions, and eventually use them to revegetate AHM sites. We will prove that our plants are AHMT with greenhouse and field trials over the two year project term. We will identify native plants that are highly desired by mine land reclamationists by using end-user surveys and canvassing our own contacts. We will have an inventory of AHMT plants for use by August 2010. Revegetation of degraded mine land will benefit society by reducing wind and water erosion from these lands,reducing human exposure and improving human health. Reclamation and revegetation will also improve wildlife habitat, improve aesthetics and recreational values, and may help endangered species. OBJECTIVES: The proposed research when successful will provide a cost-effective, relatively rapid way to produce native, locally-adapted plants which will be used to reclaim and revegetate AHM (acid soil pH and high concentrations of heavy metals)mined land sites. The plants we will culture have not been commercially propagated by any other means. Society will benefit from these innovations in the following ways: -Wind and water erosion of AHM contaminated soils will be reduced. This in turn will reduce contamination of drinking water and direct human exposure to toxic heavy metals. -Wildlife will benefit as the pre-mining plant community is re-established. -Surface and ground water contamination will be reduced. Reducing this contamination will improve the aquatic habitat for threatened and endangered aquatic species. The overall Project Goal for our SBIR Phase II project is to complete the innovation process started in Phase I. The overall innovation process can be described as taking explants from locally adapted native plants growing on AHM contaminated sites, using plant tissue culture to multiply and root them under sterile conditions, eventually producing tissue cultured AHM tolerant(AHMT) plants, then acclimating, growing to transplantable size, and overwintering them under commercial greenhouse conditions. A key part of the process is to prove that our AHMT plants have distinct advantages over their non-AHMT counterparts in both greenhouse and field trials. By non-AHMT counterpart, we mean plants of the same species, raised either from seed or vegetatively, whose origin is from similar locations but from sites which are not contaminated with heavy metals, nor which have very low soil pH. The final part of the process is to scale up production of the AHMT plants in at commercial nursery, for eventual use in reclamation and restoration projects. The following 4 objectives will accomplish our project goal: 1. Survey end-users to identify additional AHMT candidate plants; 2. Develop techniques to produce AHMT plants from tissue culture through greenhouse production; 3. Demonstrate that the AHMT plants have distinct advantages over their non-AHMT counterparts when planted on AHM soils; and 4. Produce commercially acceptable AHMT plant inventory at a commercial plant nursery. APPROACH: 1. Species Selection. SMK Plants will develop end-user surveys addressing the species needed, number needed, and when they will be needed for current and future reclamation projects. We will conduct the end-user surveys each year to identify AHMT plants with high demand for reclamation projects in Montana, Wyoming, Colorado, and Idaho. They will send these surveys out through U.S. mail, email, and will conduct some of them in person for local parties. These surveys are extremely important to obtain crucial information and to let others know about our innovative approach. They will then rank the candidate species on both potential of commercial success (i.e. how many are needed and when are they needed) and availability on AHM sites. From this ranking they will choose three species each collection season to collect and start the AHMT production process 2. Plant Propagation. This objective has 2 components, tissue culture production and nursery production, to complete the whole process of propagation to commercial size. The goal will be to complete this process for at least 3 species per year. Procedures utilized and refined in Phase I of this project will be will be implemented for the field collection, culture initiation, multiplication, and rooting of tissue cultured (tc) plants from AHM sites. The tc plants will then be tested and procedures adjusted for optimal survival and growth under greenhouse conditions. 3. Performance Trials. Greenhouse and field trials will be conducted to compare the growth and performance of the AHM selected tc plants versus non-selected plants of the same species. Greenhouse trials will involve growing the plants in pots containing soil from AHM sites for 45 days and recording performance criteria such as percent survival, fresh and dry weight, color, etc. Field trials will involve plantings on AHM sites with performance data collected over 2 growing seasons. 4. Plant Inventory. Based on tissue culture success, performance trials, and end-user surveys, 3 species with the best commercial prospects will be selected for further tissue culture production. Following the procedures used in nursery production tasks, 3,000 of the tissue cultured plants will be greenhouse acclimated, potted up, and hardened off in commercial (1qt) containers. These plants will then be ready for revegetation projects by the fall of 2010