DNA analysis of soil extracts can be used to investigate fine root depth distribution of trees

AoB Plants, Jan 2015

Understanding the root distribution of trees by soil coring is time-consuming as it requires the separation of roots from soil and classification of roots into particular size classes. This labour-intensive process can limit sample throughput and therefore sampling intensity. We investigated the use of quantitative polymerase chain reaction (qPCR) on soil DNA extractions to determine live fine root DNA density (RDD, mg DNA m−2) for mango (Mangifera indica) trees. The specificity of the qPCR was tested against DNA extracted from 10 mango cultivars and 14 weed species. All mango cultivars and no weeds were detected. Mango DNA was successfully quantified from control soil spiked with mango roots and weed species. The DNA yield of mango root sections stored in moist soil at 23–28 °C declined after 15 days to low concentrations as roots decayed, indicating that dead root materials in moist soil would not cause false-positive results. To separate large roots from samples, a root separation method for field samples was used to target the root fragments remaining in sieved (minimum 2 mm aperture) soil for RDD comparisons. Using this method we compared the seasonal RDD values of fine roots for five mango rootstock cultivars in a field trial. The mean cultivar DNA yields by depth from root fragments in the sieved soil samples had the strongest relationship (adjusted multiple R2 = 0.9307, P < 0.001) with the dry matter (g m−2) of fine (diameter <0.64 mm) roots removed from the soil by sieving. This method provides a species-specific and rapid means of comparing the distribution and concentration of live fine roots of trees in orchards using soil samples up to 500 g.

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DNA analysis of soil extracts can be used to investigate fine root depth distribution of trees

Sean L. Bithell 0 2 Lucy T. T. Tran-Nguyen 2 Mark N. Hearnden 2 Diana M. Hartley 1 Associate Editor: Astrid Volder 0 Present address: New South Wales Department of Primary Industries, Tamworth Agricultural Institute , 4 Marsden Park Rd, Tamworth, NSW 2340 , Australia 1 CSIRO Ecosystem Sciences , GPO Box 1700, Canberra, ACT 2601 , Australia 2 Plant Industries, Northern Territory Department of Primary Industry and Fisheries , GPO Box 3000, Darwin, NT 0801 , Australia Understanding the root distribution of trees by soil coring is time-consuming as it requires the separation of roots from soil and classification of roots into particular size classes. This labour-intensive process can limit sample throughput and therefore sampling intensity. We investigated the use of quantitative polymerase chain reaction (qPCR) on soil DNA extractions to determine live fine root DNA density (RDD, mg DNA m22) for mango (Mangifera indica) trees. The specificity of the qPCR was tested against DNA extracted from 10 mango cultivars and 14 weed species. All mango cultivars and no weeds were detected. Mango DNA was successfully quantified from control soil spiked with mango roots and weed species. The DNA yield of mango root sections stored in moist soil at 23 - 28 8C declined after 15 days to low concentrations as roots decayed, indicating that dead root materials in moist soil would not cause false-positive results. To separate large roots from samples, a root separation method for field samples was used to target the root fragments remaining in sieved (minimum 2 mm aperture) soil for RDD comparisons. Using this method we compared the seasonal RDD values of fine roots for five mango rootstock cultivars in a field trial. The mean cultivar DNA yields by depth from root fragments in the sieved soil samples had the strongest relationship (adjusted multiple R2 0.9307, P , 0.001) with the dry matter (g m22) of fine (diameter ,0.64 mm) roots removed from the soil by sieving. This method provides a species-specific and rapid means of comparing the distribution and concentration of live fine roots of trees in orchards using soil samples up to 500 g. Introduction Developing information on fine root and associated biomass distribution of trees is important to a number of fundamental research areas, including within season and annual growth cycles (Hendricks et al. 2006; Milchunas 2009); root longevity and carbon turnover (Eissenstat and Yanai 1997; Valverde-Barrantes et al. 2007; Goebel et al. 2011); and water or nutrient uptake (Green and Clothier 1999; Coehlo and Borges 2004; Morgan et al. 2007). Understanding of how different species or cultivars effect fine root growth (Hendrick and Pregitzer 1996), root longevity and turnover (Eissenstat and Yanai 1997; Valverde-Barrantes et al. 2007) and water or nutrient uptake (Morgan et al. 2007; da Silva et al. 2011) is also required. To improve the productivity of horticultural tree crops, information on the root traits of rootstock cultivars associated with desirable breeding objectives such as vigour control, fruit size and yield efficiency is required (Gregory et al. 2013). For fruit tree crops, knowledge of the distribution and density of fine roots is also important to optimize the efficiency of irrigation and fertigation methods (Coehlo and Borges 2004; Morgan et al. 2007). As such there is a need for new methods that allow the rapid analysis of large numbers of field samples. In addition, it would be useful if techniques to study fine roots enable live roots to be distinguished from dead roots. Especially, as the proportion of live to dead roots and the rate of root decomposition can vary among species, as found for tropical tree species (Valverde-Barrantes et al. 2007). Therefore, methods that characterize the live fine root component of tropical trees could be more meaningful than methods for the total fine root component. Such techniques will also be useful to assess the impacts of tree root diseases (Graham et al. 2013). Our study species was mango. In production regions such as the semi-arid environment of the Northern Territory (NT) of Australia, mango fruit production occurs in the dry season and irrigation is dependant on finite sources of groundwater for irrigation and fertigation (Crane et al. 1997; Pigram 2006). In live root studies of other tree species, roots were removed from soil samples by wet sieving, then separated based on colour, texture or chemical staining (Persson 1990; Hendrick and Pregitzer 1996; Schroth 2003). The use of root colour for mango is difficult as new growth is white but becomes suberized and dark-brown to black in colour (Sharma and Sharma 1988). The window to assess washed roots before they start to decay is only 48 h for refrigerated samples (Valverde-Barrantes et al. 2007) or several weeks for samples stored at 1 2 8C (Persson 1990). Roots can be stored longer for microscopic assessments when stained and stored at 5 8C in 17 % acetic acid (Liv (...truncated)


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Sean L. Bithell, Lucy T. T. Tran-Nguyen, Mark N. Hearnden, Diana M. Hartley. DNA analysis of soil extracts can be used to investigate fine root depth distribution of trees, AoB Plants, 2015, 7, DOI: 10.1093/aobpla/plu091