Recipient Organization
UNIV OF WISCONSIN
21 N PARK ST STE 6401
MADISON,WI 53715-1218
Performing Department
Agronomy
Non Technical Summary
Soybean is a legume that requires more than 300 lbs of nitrogen to produce a 60 bu/a crop. Some of this nitrogen comes from soil organic matter, but the majority of the nitrogen is produced by a symbiotic relationship with rhizobia bacteria living on the plants roots. (Lindemann and Glover, 2003; Tien et al., 2002). This bacterium, Bradyrhizobium japonicum, fixes nitrogen from the air to produce ammonium which can be used by the plant (Balatti and Pueppke, 1992). Inoculating soybean seed with B. japonicum is widely used to ensure that adequate levels of healthy bacteria are present near the seed to facilitate N fixation. In fields where native rhizobia levels are low due to lack of soybean in the rotation or environmental stress, inoculation can increase yields by 50% or more. (Duong et al., 1984; Seneviratne et al., 2000). After an initial inoculation of a field, B. japonicum will survive for many years and future inoculation will not increase yields as much as the initial one. There is a mixture of results on the efficacy of successive inoculations after initial use of B. japonicum products. Some evidence that successive inoculation can increase yields is presented by Beuerlein, 2005 and Conley and Christmas, 2006. However a number of trials did not see yield increases (Abendroth and Elmore, 2006; Pedersen, 2003; Vitosh, 1997). In another study, use of soybean inoculant raised soybean yield in 6 of 14 site-years in fields that had been in soybean rotation in a Michigan study (Schulz and Thelen, 2008). Soybean inoculants are inexpensive and fairly convenient to use. The commercial inoculant industry in Wisconsin and the Midwest is vibrant and very competitive. Newer inoculants from several of these companies contain multiple improved strains of B. japonicum and claim better nitrogen fixation efficiency leading to higher yields. Recently, inoculant manufacturers have added lipochitooligosaccharide nod factors to their inoculant products. These products are meant to elicit responses which increase infection and nodule formation (Smith et al., 2004). Inoculation of soybean, while relatively inexpensive, can be very important to the profitability of a soybean crop. Understanding the environmental conditions when inoculation should be used would greatly benefit WI soybean growers and make crop input decisions easier. Previous research has not looked at a matrix of environmental factors that might affect the performance of inoculants.
Animal Health Component
60%
Research Effort Categories
Basic
20%
Applied
60%
Developmental
20%
Goals / Objectives
Rhizobial inoculants have widely been promoted and used in Wisconsin due to our diverse rotations. It is widely known that if a field has never been in soybean or has been removed from soybean for > 4 years that inoculants are needed. In many areas of the state however, forages and small grains have been removed from the rotation and have been replaced with a soybean-corn or corn-corn-soybean rotations. Research is lacking regarding the annual need for inoculants in these "short" rotations. Our specific objectives are: 1.To develop a fast and reliable quantitative PCR assay based on real-time PCR technology and to compare its efficiency to the MPN-Plant Infection assay. 2.To determine if rhizobial inoculation is necessary the year after flooding events. 3.To quantify the effect of crop rotation and tillage on inoculant efficacy. 4.To quantify yield response of inoculants over various environmental conditions. Deliverables: The development of patentable molecular techniques to quantify soil rhizobia at the University of Wisconsin - Madison should represent a significant improvement for Wisconsin soybean growers. Growers will be then be able to make inoculation decisions rapidly (2 days vs. 5 weeks or more) and based on objective and reliable evidence. Using data collected from this series of experiments we hope to develop a prediction matrix from which growers can accurately assess the probability that an inoculant application will lead to increased yield and profitability. Lastly, we will also author a minimum of two research and two Extension publications (preliminary titles): 1.Rotation and tillage impact on soybean yield response to rhizobia 2.Flooding impact on soil rhizobia populations.
Project Methods
Objective 1. The most probable number (MPN) is the classical technique used to characterize rhizobia infection. MPN was developed by McCrady (1915) and later improved by other researchers (Cochran, 1950). In this technique axenic soybean plantlets are inoculated with diluted soil suspensions and grown for 4 weeks in a growth-chamber of a greenhouse. Plants are scored as positive or negative on the presence or absence of nodules and the MPN of rhizobia can be deduced from the classical McCrady table (Vincent, 1970). We propose to develop a quantitative PCR assay based on real-time PCR technology using the following procedure. DNA will be extracted from soil samples by lysing bacterial cells and purifying DNA from other cell and soil components of the crude extract using the SoilMasterTM DNA extraction kit (Epicenter Biotechnology). DNA will be quantified with the PicoGreen assay (Invitrogen) using a 96-well-plate fluorimeter. PCR Primers amplifying nodulation specificity genes such as nodS, nodU, nodZ, nodV, nodW and nolA will be designed using Primer3 and tested for their ability to detect specifically soybean rhizobia (Denarie et al., 1996). Once objective one is realized we will use this technique in objectives 2-4 below. For each objective soil samples will be taken at planting to quantify rhizobia populations. Rhizobia populations will be quantified according to the procedures described in Objective 1. Grain yield and end of season rhizobia populations will be quantified at R8 (physiological maturity) soybean. Objectives 2. We will identify a minimum of 3 flooded fields that were flooded for a minimum of 2 weeks in 2008. In each flooded zone we will conduct a randomized complete block design factorial experiment with 6 replications. The treatments will include an untreated check and liquid based inoculants. Objective 3. We will use the long-term rotation experiment located at Arlington WI to quantify Objective 3. This trial was established in 1983 and contains 14 different corn and soybean rotations. The experimental design is a randomized complete block split-split design with four replications. The main plot effect is tillage (conventional vs. no-till); the split-plot effect is rotation (Continuous soybean, soybean/corn rotation, 1st year soybean and corn, 2nd year soybean and corn, and third year soybean and corn); and the split-split-plot effect is inoculant treatment. Objective 4. We will use 9 sites from the Southern, Central, and North Central Region Roundup RR variety tests to field validate Objective 1 as well as quantify the effect of environment, soil type, and residual N levels on soybean yield. Across these locations, we have a variety of soil types ranging from sand to silty clay and irrigated and rainfed environments. We will select 3 high yielding soybean varieties to test at each region and each variety will receive one of three inoculant treatments: (untreated check, Optimize, or Excalibur). These inoculant treatments will be planted in a randomized complete block design with four replications using our standard plot planting and management practices.