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Methods for Experiment 172 -

Burning

Plots in field C were burned in 2005, 2006, 2007, 2008, 2009

Fencing

Experiment E001 plots were enclosed by a fence in 1982 to exclude mammalian herbivores. Galvanized welded-wire hardware cloth with 6mm x 6mm openings was buried to a depth of 84cm. Additional hardware cloth extended 60cm above the ground, and poultry netting extended to 1.8m above the ground. During the summer of 1986, aluminum flashing was placed in the ground to a depth of 12 inches. This flashing surrounded each plot, keeping the plants from spreading between plots by below-ground vegetative reproduction. The aluminum flashing was placed 12 inches from the edge of each plot. In fall 2004, herbivore exclosure fences were removed fields A, B, and D, and from all plots in field C except the following: 2, 5, 6, 7, 8, 11, 12, 13, 14, 15, 16, 17, 18, 25, 26, 29, 30, 31, 32, 37, 46, 47, 48, 50, 51, 52, and 53. These plots were designated for e172 and were enclosed in deer fences.

Layout of Plots

Plots for this experiment are 4x4 meter squares. One meter walkways separate the plots. In the table the plot number is followed by the treatment code.

Microplot Fertilization Treatments

The "microplot" fertilization treatments are used in experiments 1, 2, 4, 8, 9, 11, 23, 25, and 52. Experiments 5, 6, 28, 36, 53, 97, 98, 100 and 172 are conducted within plots that are fertilized and should be treated as part of the fertilized plot. There are nine treatment levels, assigned a numeric label 1 to 9. This is also equivalent to the letters A through I. Treatment levels 1 through 8 differ in the amount of NH₄NO₃ added. Treatment 9 is a true control and receives no nutrients. The nutrients are applied twice a year, once in early May and once in late June. An exception is experiment 25 where time of application is experimentally manipulated. Not all experiments use all possible treatments. Nutrients are given in g/m2 for each fertilization; plot area and treatments used follow the nutrient lists. Treatment 1 to 8 receive the following base nutrients:

The amount of NH₄NO₃ fertilizer added at the different treatment levels is:

Fertilizer is mixed and spread by hand twice a year, once in early May and once in late June. Each plot gets the fertilizer mixture specified by its fertilizer treatment code. These rates are added twice a year. Actual annual N addition is calculated as: 0.34%N * rate (g/m2) * 2 times/year The trace mineral stock solutions are made by adding the following grams of reagent to 1000 ml of water: CuSO4*5H2O 9.8 g ZnSO4*7H2O 22.0 g or ZnCl2 10.5 g CoCl2*6H2O 10.0 g MnCl2*4H2O 180.0 g Na2MoO4*2H2O 6.3 g H3BO3 6.0 g The trace metal solution is made by combining 120ml of each stock solution, adding 67.2g of citric acid dissolved in deionized water and adding deionized water to bring the volume to a total of 2 liters. The working solution is then autoclaved to chelate the trace metals. Some changes and/or additions occur from year to year for some experiments. These changes are listed below. Unless specific experiments are mentioned, these differences apply to all experiments in a category. In 1982, 30ml of each stock solution was combined with 16.8g citric acid, brought to a total volume of 1 liter, and autoclaved to dissolve. 1.5ml of this solution was added to 30ml of silica sand, mixed and dried. In 1983, 60ml of each stock solution was combined with 25g of EDTA brought to 1 liter, and autoclaved to dissolve. 1ml of this solution was added to 10ml of sand and added to each plot. In 1984, 60ml of each stock solution was combined with 25g of citric acid and brought to 1 liter. In 1985, the first fertilization for experiments 1 and 2, Treatment F received an extra 23 g/m2 of CaCO3 Treatment G received an extra 118.5 g/m2 of CaCO3 Treatment H received an extra 132.5 g/m2 of CaCO3 In 1985, between fertilizations for experiments 1 and 2, Treatment G received 137.5 g/m2 of CaCO3 Treatment H received 206.25 g/m2 of CaCO3 In 1985, the second fertilization for experiments 1 and 2, Treatment F received an extra 23.0 g/m2 of CaCO3 Treatment G received an extra 252.7 g/m2 of CaCO3 Treatment H received an extra 441.1 g/m2 of CaCO3 In 1985, the first fertilization for experiment 4, Treatment E received no CaCO3 Treatment G received an extra 45.5 g/m2 of CaCO3 In 1985, both fertilizations for experiments 8, 9, and 11, Treatment F received an extra 23 g/m2 of CaCO3 Treatment G received an extra 43.5 g/m2 of CaCO3 Treatment H received an extra 57.5 g/m2 of CaCO3 In 1989, lime was added to adjust pH to the following treatments in the spring: Quantities of lime stone (g/plot):

Treatment Field A Field B Field C Field D A 0 0 0 200 B 0 0 320 200 C 0 0 320 200 D 850 320 320 200 E 1000 850 1000 200 F 1300 1200 1900 200 G 1300 320 320 200 H 2100 1600 1000 20 I 0 0 0 0

Nitrogen treatment layout

lpe172 - Percent light penetration

Light Penetration

Readings were taken .5m from the front and back of each plot. The distance from the left or right edge varied from year to year. If light readings were taken before the vegetation was sampled that year, the light readings were taken in the location of the clip strip. If light readings were taken after the vegetation had been clipped, the location of the light readings was moved to one side, or the other, of the clip strip so the bare spot would not affect the readings. Usually, a set of readings were taken from the front of the plot and a second set was taken from the back of the plot. The date, time, field, plot, side (left or right), front or back, distance from the side of the plot, range value (before 1988) and the display reading were all recorded for each measurement. If there was a gopher mound where the light readings should be taken, the sensor was moved in from the edge of the plot until it is no longer over a gopher mound. This was then recorded on the data sheet.

Light Penetration

From 1982 to 1988, light meter readings were taken using a Li-Cor, Inc. Integrating Quantum/Radiometer/Photometer, model LI-188B. Two people were needed to take light readings with this system. One would hold the control box and record data and the other would hold the light sensor. The batteries would be checked before starting each session. If the batteries were OK, they would proceed to take readings. The sensor was connected to the control box by a relatively short cord, so the two people taking readings were required to stay close together. When taking readings, it was necessary to get a range value for each light value entered. If the integrating time of 1 second was not sufficient, it was increased to 10 seconds. This was also recorded. In 1989, two new light meters were acquired. These are SF Sunfleck Ceptometers, model SF-40 (40cm probe). They were purchased from Decagon Devices, Inc., P.O. Box 835, Pullman, WA 99163. One person can easily handle a ceptometer alone. This makes is possible for three people to get the readings done more quickly and easily. One person records data while the other two take readings from the plots simultaneously. No range values are needed. To take readings a person needs to select function #1 (PAR readings), position the probe (see below) and press ``A' (read value). More than one reading can be taken and then averaged by pressing a certain sequence of letters (A, A, B, B, A). Measurements are taken within a 4 hour period, 2 hours on either side of solar noon. (Solar noon is half way between sunrise and sunset; it is not 1200 hours). Solar noon is at 1315 hours, Central Daylight Time. Samples are taken between 1115 hours and 1515 hours. Measurements are not taken when the plot being sampled is shaded. Light readings are done when the sky is clear, whenever possible. If a cloud passes over the plot being sampled, assistants wait for the cloud to pass before taking the readings. If the sky is mostly cloudy, light meter readings are not taken. Two measurements are taken in each plot. Each measurement consists of one reading above the vegetation and a second reading at ground level. Both values are taken to get the percent of sunlight above the vegetation that reaches ground level. In taking the above vegetation reading, the sensor must be kept level, held high above all vegetation, kept out of the shade (of plants and people) and it must be clean. When taking the below vegetation reading, at ground level, the sensor must be kept level, out of the soil and out of the shade created by people. Light Data Transformations: Light readings are transformed to obtain percent light penetration which represents the percent of light above the vegetation that reaches the ground surface. In cases where the experiment involves shading, another variable is computed to reflect the percent of sun light that reaches above the canopy. This variable is called light available. In the case of absence of artificial shades, the latter is set to 1. percent light penetration = ( Light below canopy / Light above canopy ). percent light available = ( Light below shade / Light above shade ).

ple172 - Plant aboveground biomass data

Above ground sampling

Aboveground vegetation is sampled annually. Sampling is done the same way in each of the four fields, A, B, C, and D. Field A is sampled first, usually in early July. Field B is sampled second, usually in late July or early August. Field C is sampled third, usually in mid-August. Field D is sampled last, usually in late August. These sampling times were chosen to correspond with the usual time of peak biomass in each of the fields. Each year, the sample strip is located in a different place. A 10cm x 3m strip of vegetation is clipped out of each plot. Aboveground vegetation is collected in narrow strips to limit the effect of the sampling. Benches are used to avoid trampling the surrounding area. A 3m long fiberglass pole is placed along the ground next to the appropriate site of the present year's sample strip. This is used as a guide to keep the 3m long sample strip straight. Vegetation is cut using a Sears/Craftsman Extra Heavy Duty Cordless Electric Grass Shears, model # 240.79672. These clippers have blades which are 10cm wide, making it easy to clip a strip of that width. The clippers are powered by 6-volt motorcycle batteries. All vegetation that is rooted in the 10cm wide strip is cut 1-2cm from the soil surface. The vegetation and litter that was in the strip is collected. Care is taken to clip and collect only that material which is actually in the 10cm wide strip, not entire plants that may lay across the strip. If there is a gopher mound in the plot where the clip strip is, the strip is moved in towards the middle of the plot until it is no longer over a gopher mound. The new position of the clip strip is then recorded. Year Clipped Area 1982 Clipped 0.25 to 0.35m from left, 0.5 to 3.5m from bottom 1982 Clipped 3.65 to 3.75m from left, 0.5 to 3.5m from bottom 1983 Clipped 0.5 to 0.6m from left, 0.5 to 3.5m from bottom 1984 Clipped 3.4 to 3.5m from left, 0.5 to 3.5m from bottom 1985 Clipped 0.75 to 0.85m from left, 0.5 to 3.5m from bottom 1986 Clipped 3.15 to 3.25m from left, 0.5 to 3.5m from bottom 1987 Clipped 0.9 to 1m from left, 0.5 to 3.5m from bottom 1988 Clipped 3 to 3.1m from left, 0.5 to 3.5m from bottom 1989 Clipped 0.35 to 0.45m from left, 0.5 to 3.5m from bottom 1990 Clipped 3.55 to 3.65m from left, 0.5 to 3.5m from bottom 1991 Clipped 0.6 to 0.7m from left, 0.5 to 3.5m from bottom 1992 Clipped 3.3 to 3.4m from left, 0.5 to 3.5m from bottom 1993 Clipped 3.65 to 3.75m from left, 0.5 to 3.5m from bottom 1994 Clipped 0.25 to 0.35m from left, 0.5 to 3.5m from bottom 1995 Clipped 3.4 to 3.5m from left, 0.5 to 3.5m from bottom 1996 Clipped 0.5 to 0.6m from left, 0.5 to 3.5m from bottom 1997 Clipped 3.15 to 3.25m from left, 0.5 to 3.5m from bottom 1998 Clipped 0.75 to 0.85m from left, 0.5 to 3.5m from bottom 1999 Clipped 3 to 3.1m from left, 0.5 to 3.5m from bottom 2000 Clipped 0.9 to 1m from left, 0.5 to 3.5m from bottom 2001 Clipped 1.35 to 1.45m from left, 0.5 to 3.5m from bottom 2002 Clipped 3.65 to 3.75m from left, 0.5 to 3.5m from bottom 2003 Clipped 1.80 to 1.90m from left, 0.5 to 3.5m from bottom 2004 Clipped 1.90 to 1.80m from right, 0.5 to 3.5m from bottom 2005 Clipped 1.20 to 1.30m from left, 0.5 to 3.5m from bottom 2006 Clipped 1.60 to 1.50m from right, 0.5 to 3.5m from bottom Sampling Discrepancies in E001-E172 Field C Year Sampling Discrepancy 1982 none 1983 Plot 15 clipped at 0.60-0.70m from left side to avoid a gopher mound 1984 Plot 31 to Plot 49 clipped at 0.50-0.60m from left side 1985 none 1986 Plot 43 clipped at 0.75-0.85m from left side, Plot 45 clipped at 0.75-0.85m from left side, Plot 46 clipped at 0.75-0.85m from left side 1987 none 1988 Plot 12 clipped at 3.55-3.65cm from left side 1989 none 1990 none 1991 none 1992 none 1993 none 1994 none 1995 none 1996 none 1997 none 1998 none 1999 none 2000 none 2001 Plot 50 clipped 2.55-2.65m from left due to gopher disturbance 2002 none 2003 none 2004 none 2005 Plots 34 clipped at 0.20-0.10m from right side 2006 none

rbe172 - Root biomass data

Root biomass sampling

Root samples are obtained using a two inch diameter corer. Three core samples of depth 0-30cm are taken from the area that has been clipped for aboveground biomass within a few days after clipping. The roots and associated crowns are extracted from the sample by washing with water above a screen. Any extraneous non-root material is removed. The cleaned roots are dried and weighted. In 1994 samples were taken from all plots in the experiment. Three cores are taken from each plot using the two inch diameter corer and combined into one sample. Cores are taken to a depth of 15cm.