Harvard Forest

Data Archive

HF297

Foliar and Soil Chemistry at Harvard Forest Chronic Nitrogen Amendment Experiment 1995-2009

Related Publications

Data

• hf297-01: pine site data (preview)
• hf297-02: hardwood site oak (preview)
• hf297-03: hardwood site maple (preview)
• hf297-04: soil chemistry and metabolites (preview)
• hf297-05: microbial diversity (preview)
• hf297-06: OTU rep sequences
• hf297-07: gene bank accession numbers

Overview

• Lead: Rakesh Minocha, Stephanie Long, Swathi Turlapati
• Investigators: Subhash Minocha
• Contact: Information Manager
• Start date: 1995
• End date: 2009
• Status: complete
• Location: Prospect Hill Tract (Harvard Forest)
• Latitude: +42.54 degrees
• Longitude: -72.18 degrees
• Elevation: 385 meter
• Datum: WGS84
• Taxa: Acer rubrum (red maple), Pinus resinosa (red pine), Quercus rubra (red oak), Quercus velutina (black oak)
• Release date: 2023
• Language: English
• EML file: knb-lter-hfr.297.5
• DOI: digital object identifier
• EDI: data package
• DataONE: data package
• Related links:
• Chronic Nitrogen Amendment Experiment at Harvard Forest since 1988
• Study type: long-term measurement
• Research topic: large experiments and permanent plot studies; physiological ecology, population dynamics and species interactions
• LTER core area: population studies, mineral cycling, disturbance patterns
• Keywords: amino acids, bacteria, chlorophyll, foliage, genomics, microbes, soil solution chemistry
• Abstract:

  The aim of the chronic N study at Harvard Forest is to increase our understanding of ecosystem nitrogen dynamics in response to elevated nitrogen inputs. In recent, nitrogen deposition in the Northeastern United States has been 10 to 20 times above historic background levels which could possibly saturate the retention capacity of a forest ecosystem. Long-term elevated N deposition typically leads to an increase in the concentration of total foliar N, with or without similar changes in the important base elements such as Ca, Mg and K. This increase in leaf N content also leads to significant shifts in the internal partitioning of N within the leaf. For example, in conifers, N deposition has been shown to significantly increase leaf N present in the form of free amino acids such as arginine. Little is known about N partitioning for hardwoods under these conditions. These changes in N partitioning are possibly connected to leaf function. The present study was conducted to experimentally test whether the alterations in N partitioning do occur due to long-term N deposition and if so do they have a positive or a negative effect on photosynthetic capacity and biomass production. A possible decoupling of the relationship between foliar N and photosynthetic rate may occurs under these conditions.

  The treatment plots used in this study are part of the Chronic Nitrogen Amendment Study at the Harvard Forest LTER site (42.5°N, 72°W). The site has a temperate climate with monthly temperatures ranging from -7°C in January to 20°C in July. Average annual precipitation is 110 cm (http://harvardforest.fas.harvard.edu). The site averages approximately 8 kg ha-1 year-1 of total N deposition. As reported earlier, the land-use history of the pine and hardwood stands used in this study is very different. Two adjacent stands were chosen for the study: an even-aged red pine (Pinus resinosa Ait.) stand and a 50-year-old mixed hardwood stand that had regenerated naturally after clearcutting in approximately 1945. The hardwood stand is dominated by black and red oak (Quercus velutina Lam.; Q. rubra L.) with significant amounts of black birch (Betula lenta L.), red maple (Acer rubrum L.) and American beech (Fagus grandifolia Ehrh.). The pine site was pastured until 1920, and a red pine (Pinus resinosa Ait.) plantation was established on this site in 1926. Fire destroyed some of the trees in 1957. The hardwood stand has no agricultural land-use history but was substantially damaged by a hurricane in 1938, and was harvested for firewood during 1942-44. In 1988, 30 m x 30 m plots were established in each of the two adjacent stands, red pine and a mixed deciduous each. Starting in 1989, while one plot served as control, six equal doses of NH4NO3 were applied to Low N (50 kg N ha-1 year-1) and High N (150 kg N ha-1 year-1) plots of each stand from May through September each year.

• Methods:

  Fertilization

  Fertilizer additions of NH4NO3 and Na2SO4 began in 1988 as six equal applications over the growing season (May - Sept). Fertilizer is weighed, mixed with 20 liters of water (equivalent to 0.002 cm rainfall) and applied using a backpack sprayer. Na2SO4 applications were discontinued after the 1998 growing season. The hardwood and pine stands each contain four plots, with application rates as follows: Control, Low = 5gN/m^2 per year, High = 15gN/m^2 per year, N+S = 5gN/m^2 + 7.4gS/m^2 per year. Plot designations are P or H followed by C, L, H, NS for control, and low, high N+ S, respectively. Upon request, more detailed information is available on the exact applications rates based on fertilizer sample analysis.

  Foliar Collections

  Foliar collections were done on the same trees at the same time as the Aber and Magill group.

  Photosynthesis and Related Measurements in 2000

  Refer to: Bauer, G. A., F. A. Bazzaz, R. Minocha, S. Long, A. Magill, J. Aber, and G. M. Berntson. 2004. Effects of chronic N additions on tissue chemistry, photosynthetic capacity, and carbon sequestration potential of a red pine (Pinus resinosa Ait.) stand in the NE United States. Forest Ecology and Management 196:173-186 for detail.

  Other methods are described in: Minocha, R., S. A. Turlapati, S. Long, W. H. McDowell, and S. C. Minocha. 2015. Long-Term Trends of Changes in Pine and Oak Foliar Nitrogen Metabolism in Response to Chronic Nitrogen Amendments at the Harvard Forest, MA. Tree Physiology 35:894-909.

  Methods for 1995-2008 Metabolic Analyses

  Samples of red pine and black and red oak (Quercus velutina Lam.; Q. rubra L., hereafter referred to only as oak) were collected in July or August 1995, 1996, 1997, 2002, 2005 and 2008 from mid to upper canopy of mature trees by shooting down small branches of asymptomatic trees with a shotgun. The number of trees sampled varied with year and tree species. Each year trees were selected randomly from each treatment plot. In general, with the exception of 2005 for pine and 1997 for oak where sample sizes were five, ten or more trees were sampled for other years. More detailed information for sample size for each year is provided in the legend of each figure. All samples were processed as described in Minocha et al. (2000). Briefly, for each sample, a pool of approximately 500 mg fresh weight (FW) of foliage was collected from a single branch from each tree. Current year needles of pine were cut with scissors into 1-2 mm pieces, and 4 mm disks were punched from oak leaves with a paper punch carefully excluding major veins. The clippings were mixed and a sample (~200 mg FW) was placed in a pre-weighed 2 mL microfuge tube with one mL of 5% perchloric acid (PCA). All samples were immediately placed on ice for transport to the laboratory and stored at -20o C until further analysis. The tubes with tissue sample and PCA were weighed, frozen and thawed 3 times and then centrifuged at 13,000 xg for 10 min. The resulting supernatant was used for analyses of PCA-extractable (free) PAs and AAs, and soluble inorganic elements. For each analysis, all extracts were analyzed individually without pooling.

  For analyses of PAs and AAs, the supernatant of the PCA extracted samples was subjected to dansylation and quantitation by HPLC (PerkinElmer Inc., Waltham MA, USA) according to previously published protocols (Minocha et al. 2000; Minocha and Long 2004b). The reaction was terminated using L-asparagine (50 L of 20 mg mL-1 in water) rather than alanine as described in the original method. In using this protocol the separation of Arg and threonine (Thr) was not always complete; for quantitation, the peak areas of these two AAs were added together to formulate a combined calibration curve.

  Inorganic ions (soluble in 5% PCA) were quantified using a simultaneous axial Inductively Coupled Plasma emission spectrophotometer (Vista CCD, Varian Inc., Palo Alto, CA, USA) and Vista Pro software (Version 4.0), following appropriate dilutions with deionized water.

  Sapwood Plugs

  In 2005 and 2008, sapwood plugs from actively growing clear (no stained or rotten wood) sapwood were extracted from each tree using a drill and an increment hammer. Samples were taken at breast height, avoiding areas with obvious injury to the stem. Three sapwood plugs for oak and 4 for pine were collected from different faces of each tree. A drill was used to remove the bark and the cambial layer. A dead blow hammer was then used to drive the increment hammer into the exposed sapwood. A plunger was used to extract the sapwood plug into labeled Ziploc bags. Plugs were approximately 1.5 cm in length. Prior to processing, each plug was cut into 4-6 equal segments. After removing any extraneous materials such as bark or discolored wood, samples (~600 mg FW) were transferred into pre-weighed and labeled microfuge tubes. Following the addition of one mL of 5% PCA, samples were stored on ice in the field and frozen at -20oC in the lab until further analysis. Sapwood samples were processed for PA, AA and ion analysis as described above for foliage.

  Soil Solution Chemistry

  Details of the installation of Zero-Tension Lysimeters (ZTL) and soil solution sample collection are described in Minocha et al. (2000) and references therein. Briefly, 5 polyethylene ZTL’s were installed per treatment plot. Soil solution was collected after major rain events. All 5 samples per plot were pooled prior to analyses. Over the course of each year, there were ~50 collections from each plot. Samples were transported on ice to the laboratory and filtered through pre-combusted Whatman GF/F glass fiber filters (Whatman Inc., Clifton, NJ) within 36 h of collection before freezing. These solutions were analyzed for inorganic elements using ICP as described above.

  Annual Mortality Data

  Mortality numbers were calculated each year as percent of dead trees compared to the number of live trees at the beginning of the study in 1988. In order to keep the number of trees constant for mortality calculations for the entire duration of the study, those trees that were less than 5 cm DBH in size at the beginning of the study (1988) but grew to more than 5 cm by summer 2008 were included in the total number of trees in inventory of 1988-2008.

  Tree Inventory

  This part was done by Aber and Ollinger groups. All trees above 5cm Diameter Breast Height (DBH) were numbered and measured in 1988. In subsequent years, trees previously too small were tagged and measured. In this dataset, we report tree-level DBH measurements. Tree data is reported by subplot - numbered [a-f][[1-6], for a total of 36 5x5m subplots per plot. Note that in some years, some subplots were not sampled (HC-F1, PL-F[1-6]). For mortality tables see, Minocha et al. 2015.

  Zero Tension Lysimeters

  ZTL's are gravity fed gravel and glass bead filled collection units (0.3 to 1.5cm) located beneath the organic horizon with a tube draining approximately 154 cm2 into a 1 liter bottle. Samples were collected within 1 week following a rain event, weighed, filtered through a 0.7um filter and frozen prior to analysis. Base cations were measured using either ICP (1998 and later) or DCP (1997 and earlier) For more details see Minocha et al. 2000 and 2015.

  Soil Microbial Diversity and Soil Chemistry in 2009

  In September 2009, soil cores were collected from five randomly selected subplots within each treatment plot using a soil corer (7.5 cm diameter) to a depth of approximately 15 cm. For each soil core, the upper, dark brown organic horizon (Org) was separated from the lower, light brown mineral horizon (Min). A total of 30 soil samples (5 cores per plot x 2 horizons x 3 treatments) were collected and brought to the laboratory on ice in polyethylene bags. Each sample was given a designation consisting of the treatment name-subplot name-soil horizon. The samples were sieved (2-mm pore size) to remove roots and stones and stored at -20 °C until further use.

  Soil Chemical Analyses

  Air-dried soil samples (20-40 g) were sent to the Soil Testing Service Laboratory at the University of Maine, Orono, ME, USA (http://anlab.umesci.maine.edu) for chemical analyses. The total N and C contents were measured by combustion analysis at 1,350 ºC. Exchangeable P, Al and base cations were extracted with 1 M NH4Cl at a ratio of 2 g of organic soil or 5g of mineral soil to 100 mL of extraction solution (Blume et al., 1990). After shaking for one hour, the extracts were vacuum filtered (Whatman # 42, Whatman Inc., Clifton, NJ, USA) and analyzed using flame emission spectroscopy for K and Na and plasma emission spectroscopy for Ca, Mg, and Al. The exchangeable acidity was measured in 1 M KCl extracts using end-point titration. The percentage of soil organic matter in the oven-dried samples was determined using loss-on-ignition (LOI) for 12 h at 550 ºC with a muffle furnace. The effective cation-exchange capacity (ECEC) was calculated as the sum of the exchangeable base cations (Ca, Mg, K, and Na) plus the exchangeable acidity. For more details please refer to Turlapati et al. 2013.

  DNA Isolation, PCR Amplification and Pyrosequencing

  Total DNA was isolated from 0.5 g of each soil sample using the PowerSoil® DNA isolation kit (MO-BIO Laboratories, Carlsbad, CA, USA) following the manufacturer’s instructions, with the minor modification that included bead beating only for 7 min with vortex adaptor to minimize DNA shearing. The obtained DNA was quantified and examined for purity (A260:A280 ratio between 1.6-1.8 and A260:A230 ratio between 2.0-2.2) with a NanoDrop spectrophotometer (Thermo-Fisher Scientific, Waltham, MA, USA). In preparation for pyrosequencing, universal primers with 30 different barcodes (10 bp, one for each of the 30 soil samples) were designed to amplify a 433-bp fragment of the hypervariable region (V6-V8) of the bacterial 16S rRNA gene from the soil samples. The primers used were F968 (5’AA CGC GAA GAACCT TAC3’) and R1401-1a (5’CGG TGT GTA CAA GGC CCG GGA ACG3’) as described in Brons and van Elsas (2008). The amplification reactions were conducted in triplicate using Phusion® Taq Master Mix (New England Biolabs, Ipswich, MA, USA) with 50 ng of template DNA in a final volume of 50 µL. The reactions were performed in a PTC-100® Programmable Thermal Cycler (MJ Research, Inc., Waltham, MA, USA) with the following conditions: an initial denaturation at 95 ºC for five min, followed by 20 cycles of denaturation at 95 ºC for 30 sec, annealing at 61 ºC for 30 sec, and extension at 72 ºC for 45 sec, with a final extension at 72 ºC for 10 min. The triplicate reaction products (amplicons) from each soil sample were pooled for sequencing (Margulies et al., 2005). The pooled PCR products were purified using a DNA purification kit (Zymo Research, Irvine, CA, USA) and subjected to further cleaning via the Agencourt® AMPure® XP Bead Purification method (Agencourt Bioscience Corporation, Beverly, MA, USA) to remove fragments less than 100 bp. The quality of the PCR products was evaluated in an Agilent 2100 Bioanalyzer using the DNA 1000 LabChip (Agilent Technologies, Palo Alto, CA, USA). The 30 bar-coded samples were pooled in equimolar quantities and processed for sequencing in a full picotiter plate (Roche 454 GS-FLX Titanium System) at the University of Illinois, USA (www.biotec.illinois.edu/centers/Keck/Highthroughput/).

  Data Processing

  The sequencing data were processed using the Quantitative Insights into Microbial Ecology (QIIME) toolkit Version 1.4.0 with default settings for most of the steps, except where specified. The pipeline used for data processing is described in Turlapati et al. 2013.or oligotyping as described in Turlapati et al. 2015. The OTU representative sequences were submitted to NCBI Genbank and were assigned accession numbers JQ049082-JQ060105.

• Organization: Harvard Forest. 324 North Main Street, Petersham, MA 01366, USA. Phone (978) 724-3302. Fax (978) 724-3595.

• Project: The Harvard Forest Long-Term Ecological Research (LTER) program examines ecological dynamics in the New England region resulting from natural disturbances, environmental change, and human impacts. (ROR).

• Funding: National Science Foundation LTER grants: DEB-8811764, DEB-9411975, DEB-0080592, DEB-0620443, DEB-1237491, DEB-1832210.

• Use: This dataset is released to the public under Creative Commons CC0 1.0 (No Rights Reserved). Please keep the dataset creators informed of any plans to use the dataset. Consultation with the original investigators is strongly encouraged. Publications and data products that make use of the dataset should include proper acknowledgement.

• License: Creative Commons Zero v1.0 Universal (CC0-1.0)

• Citation: Minocha R, Long S, Turlapati S. 2023. Foliar and Soil Chemistry at Harvard Forest Chronic Nitrogen Amendment Experiment 1995-2009. Harvard Forest Data Archive: HF297 (v.5). Environmental Data Initiative: https://doi.org/10.6073/pasta/299c2e85eef1ab360a469e85188c61a6.

Detailed Metadata

hf297-01: pine site data

1. sitename: collection site, study name
2. year: year of sample collection
3. species: tree species
   • PIRE: Pinus resinosa Aiton (red pine)
   • PINE: pine
4. subplot: subplot designation within site
5. treetag: tree tag number
6. treatment: treatment
   • PC: pine control
   • PL: pine low N
   • PH: pine high N
7. tissue: sample type
8. exch.ca: five percent PCA soluble Ca, by ICP (unit: micromolePerGram / missing value: NA)
9. exch.k: five percent PCA soluble K, by ICP (unit: micromolePerGram / missing value: NA)
10. exch.mg: five percent PCA soluble Mg, by ICP (unit: micromolePerGram / missing value: NA)
11. exch.mn: five percent PCA soluble Mn (unit: micromolePerGram / missing value: NA)
12. exch.p: five percent PCA soluble P, by ICP (unit: micromolePerGram / missing value: NA)
13. exch.al: five percent PCA soluble Al, by ICP (unit: micromolePerGram / missing value: NA)
14. exch.fe: five percent PCA soluble Fe, by ICP (unit: micromolePerGram / missing value: NA)
15. exch.na: five percent PCA soluble Na, by ICP (unit: micromolePerGram / missing value: NA)
16. exch.zn: five percent PCA soluble Zn, by ICP (unit: micromolePerGram / missing value: NA)
17. tot.chloro: total chlorophyll by spectrophotometer (unit: microgramPerGram / missing value: NA)
18. chloro.a: total chlorophyll by spectrophotometer (unit: microgramPerGram / missing value: NA)
19. chloro.b: total chlorophyll by spectrophotometer (unit: microgramPerGram / missing value: NA)
20. chloro.a.b: total chlorophyll by spectrophotometer (unit: microgramPerGram / missing value: NA)
21. sol.proteins: soluble proteins by spectrophotometer (unit: milligramPerGram / missing value: NA)
22. put: five percent PCA soluble putrescine by HPLC (unit: nanomolePerGram / missing value: NA)
23. spd: five percent PCA soluble spermidine by HPLC (unit: nanomolePerGram / missing value: NA)
24. spm: five percent PCA soluble spermine by HPLC (unit: nanomolePerGram / missing value: NA)
25. spd.put: ratio of spermidine to putrescine (unit: dimensionless / missing value: NA)
26. asp: five percent PCA soluble aspartic acid by HPLC (unit: nanomolePerGram / missing value: NA)
27. glu: five percent PCA soluble glutamic acid by HPLC (unit: nanomolePerGram / missing value: NA)
28. gln: five percent PCA soluble glutamine by HPLC (unit: nanomolePerGram / missing value: NA)
29. ser: five percent PCA soluble serine by HPLC (unit: nanomolePerGram / missing value: NA)
30. arg: five percent PCA soluble arginine by HPLC (unit: nanomolePerGram / missing value: NA)
31. thr: five percent PCA soluble threonine by HPLC (unit: nanomolePerGram / missing value: NA)
32. gly: five percent PCA soluble glycine by HPLC (unit: nanomolePerGram / missing value: NA)
33. arg.thr: five percent PCA soluble arginine+threonine by HPLC (system was unable to separate these two amino acids) (unit: nanomolePerGram / missing value: NA)
34. arg.thr.gly: five percent PCA soluble arginine+threonine+glycine by HPLC (system was unable to separate these three amino acids) (unit: nanomolePerGram / missing value: NA)
35. ala: five percent PCA soluble alanine by HPLC (unit: nanomolePerGram / missing value: NA)
36. pro: five percent PCA soluble proline by HPLC (unit: nanomolePerGram / missing value: NA)
37. gaba: five percent PCA soluble g-aminobutyric acid by HPLC (unit: nanomolePerGram / missing value: NA)
38. val: five percent PCA soluble valine by HPLC (unit: nanomolePerGram / missing value: NA)
39. met: five percent PCA soluble methionine (unit: nanomolePerGram / missing value: NA)
40. ile: five percent PCA soluble isoleucine by HPLC (unit: nanomolePerGram / missing value: NA)
41. leu: five percent PCA soluble leucine by HPLC (unit: nanomolePerGram / missing value: NA)
42. trp: five percent PCA soluble tryptophan by HPLC (unit: nanomolePerGram / missing value: NA)
43. phe: five percent PCA soluble phenylalanine by HPLC (unit: nanomolePerGram / missing value: NA)
44. ile.leu: five percent PCA soluble isoluecine+leucine by HPLC (system was unable to separate these two amino acids) (unit: nanomolePerGram / missing value: NA)
45. leu.trp: five percent PCA soluble isoluecine+tryptophan by HPLC (system was unable to separate these two amino acids) (unit: nanomolePerGram / missing value: NA)
46. trp.phe: five percent PCA soluble tryptophan+phenylalanine by HPLC (system was unable to separate these two amino acids) (unit: nanomolePerGram / missing value: NA)
47. cys: five percent PCA soluble cystine by HPLC (unit: nanomolePerGram / missing value: NA)
48. orn: five percent PCA soluble ornithine by HPLC (unit: nanomolePerGram / missing value: NA)
49. lys: five percent PCA soluble lysine by HPLC (unit: nanomolePerGram / missing value: NA)
50. his: five percent PCA soluble histidine by HPLC (unit: nanomolePerGram / missing value: NA)

hf297-02: hardwood site oak

1. sitename: collection site, study name
2. year: year of sample collection
3. species: tree species
   • QURU/QUVE: Quercus rubra L. (red oak)/Quercus veutina Lam. (black oak)
   • HARDWOOD: hardwood
4. subplot: subplot designation within site
5. treetag: tree tag number
6. treatment: treatment
   • HC: hardwood control
   • HL: hardwood low N
   • HH: hardwood high N
7. tissue: sample type
8. exch.ca: five percent PCA soluble Ca, by ICP (unit: micromolePerGram / missing value: NA)
9. exch.k: five percent PCA soluble K, by ICP (unit: micromolePerGram / missing value: NA)
10. exch.mg: five percent PCA soluble Mg, by ICP (unit: micromolePerGram / missing value: NA)
11. exch.mn: five percent PCA soluble Mn, by ICP (unit: micromolePerGram / missing value: NA)
12. exch.p: five percent PCA soluble P, by ICP (unit: micromolePerGram / missing value: NA)
13. exch.al: five percent PCA soluble Al, by ICP (unit: micromolePerGram / missing value: NA)
14. exch.fe: five percent PCA soluble Fe, by ICP (unit: micromolePerGram / missing value: NA)
15. exch.na: five percent PCA soluble Na, by ICP (unit: micromolePerGram / missing value: NA)
16. exch.zn: five percent PCA soluble Zn, by ICP (unit: micromolePerGram / missing value: NA)
17. tot.chloro: total chlorophyll by spectrophotometer (unit: microgramPerGram / missing value: NA)
18. chloro.a: total chlorophyll by spectrophotometer (unit: microgramPerGram / missing value: NA)
19. chloro.b: total chlorophyll by spectrophotometer (unit: microgramPerGram / missing value: NA)
20. chloro.a.b: total chlorophyll by spectrophotometer (unit: microgramPerGram / missing value: NA)
21. sol.proteins: soluble proteins by spectrophotometer (unit: milligramPerGram / missing value: NA)
22. put: five percent PCA soluble putrescine by HPLC (unit: nanomolePerGram / missing value: NA)
23. spd: five percent PCA soluble spermidine by HPLC (unit: nanomolePerGram / missing value: NA)
24. spm: five percent PCA soluble spermine by HPLC (unit: nanomolePerGram / missing value: NA)
25. spd.put: ratio of spermidine to putrescine (unit: dimensionless / missing value: NA)
26. asp: five percent PCA soluble aspartic acid by HPLC (unit: nanomolePerGram / missing value: NA)
27. glu: five percent PCA soluble glutamic acid by HPLC (unit: nanomolePerGram / missing value: NA)
28. gln: five percent PCA soluble glutamine by HPLC (unit: nanomolePerGram / missing value: NA)
29. ser: five percent PCA soluble serine by HPLC (unit: nanomolePerGram / missing value: NA)
30. arg: five percent PCA soluble arginine by HPLC (unit: nanomolePerGram / missing value: NA)
31. thr: five percent PCA soluble threonine by HPLC (unit: nanomolePerGram / missing value: NA)
32. gly: five percent PCA soluble glycine by HPLC (unit: nanomolePerGram / missing value: NA)
33. arg.thr: five percent PCA soluble arginine+threonine by HPLC (system was unable to separate these two amino acids) (unit: nanomolePerGram / missing value: NA)
34. arg.thr.gly: five percent PCA soluble arginine+threonine+glycine by HPLC (system was unable to separate these three amino acids) (unit: nanomolePerGram / missing value: NA)
35. ala: five percent PCA soluble alanine by HPLC (unit: nanomolePerGram / missing value: NA)
36. pro: five percent PCA soluble proline by HPLC (unit: nanomolePerGram / missing value: NA)
37. gaba: five percent PCA soluble g-aminobutyric acid by HPLC (unit: nanomolePerGram / missing value: NA)
38. val: five percent PCA soluble valine by HPLC (unit: nanomolePerGram / missing value: NA)
39. met: five percent PCA soluble methionine by HPLC (unit: nanomolePerGram / missing value: NA)
40. ile: five percent PCA soluble isoleucine by HPLC (unit: nanomolePerGram / missing value: NA)
41. leu: five percent PCA soluble leucine by HPLC (unit: nanomolePerGram / missing value: NA)
42. trp: five percent PCA soluble tryptophan by HPLC (unit: nanomolePerGram / missing value: NA)
43. phe: five percent PCA soluble phenylalanine by HPLC (unit: nanomolePerGram / missing value: NA)
44. ile.leu: five percent PCA soluble isoluecine+leucine by HPLC (system was unable to separate these two amino acids) (unit: nanomolePerGram / missing value: NA)
45. leu.trp: five percent PCA soluble isoluecine+tryptophan by HPLC (system was unable to separate these two amino acids) (unit: nanomolePerGram / missing value: NA)
46. trp.phe: five percent PCA soluble tryptophan+phenylalanine by HPLC (system was unable to separate these two amino acids) (unit: nanomolePerGram / missing value: NA)
47. cys: five percent PCA soluble cystine by HPLC (unit: nanomolePerGram / missing value: NA)
48. orn: five percent PCA soluble ornithine by HPLC (unit: nanomolePerGram / missing value: NA)
49. lys: five percent PCA soluble lysine by HPLC (unit: nanomolePerGram / missing value: NA)
50. his: five percent PCA soluble histidine by HPLC (unit: nanomolePerGram / missing value: NA)

hf297-03: hardwood site maple

1. sitename: collection site, study name
2. year: year of sample collection
3. species: tree species
   • ACRU: Acer rubrum L. (red maple)
   • HARDWOOD: hardwood
4. subplot: subplot designation within site
5. treetag: tree tag number
6. treatment: treatment
   • HC: hardwood control
   • HL: hardwood low N
   • HH: hardwood high N
7. tissue: sample type
8. exch.ca: five percent PCA soluble Ca, by ICP (unit: micromolePerGram / missing value: NA)
9. exch.k: five percent PCA soluble K, by ICP (unit: micromolePerGram / missing value: NA)
10. exch.mg: five percent PCA soluble Mg, by ICP (unit: micromolePerGram / missing value: NA)
11. exch.mn: five percent PCA soluble Mn, by ICP (unit: micromolePerGram / missing value: NA)
12. exch.p: five percent PCA soluble P, by ICP (unit: micromolePerGram / missing value: NA)
13. exch.al: five percent PCA soluble Al, by ICP (unit: nanomolePerGram / missing value: NA)
14. exch.fe: five percent PCA soluble Fe, by ICP (unit: micromolePerGram / missing value: NA)
15. exch.na: five percent PCA soluble Na, by ICP (unit: micromolePerGram / missing value: NA)
16. exch.zn: five percent PCA soluble Zn, by ICP (unit: micromolePerGram / missing value: NA)
17. tot.chloro: total chlorophyll by spectrophotometer (unit: microgramsPerGram / missing value: NA)
18. chloro.a: total chlorophyll by spectrophotometer (unit: microgramsPerGram / missing value: NA)
19. chloro.b: total chlorophyll by spectrophotometer (unit: microgramsPerGram / missing value: NA)
20. chloro.a.b: total chlorophyll by spectrophotometer (unit: microgramsPerGram / missing value: NA)
21. sol.proteins: soluble proteins by spectrophotometer (unit: microgramsPerGram / missing value: NA)
22. put: five percent PCA soluble putrescine by HPLC (unit: nanomolePerGram / missing value: NA)
23. spd: five percent PCA soluble spermidine by HPLC (unit: nanomolePerGram / missing value: NA)
24. spm: five percent PCA soluble spermine by HPLC (unit: nanomolePerGram / missing value: NA)
25. spd.put: ratio of spermidine to putrescine (unit: dimensionless / missing value: NA)
26. asp: five percent PCA soluble aspartic acid by HPLC (unit: nanomolePerGram / missing value: NA)
27. glu: five percent PCA soluble glutamic acid by HPLC (unit: nanomolePerGram / missing value: NA)
28. gln: five percent PCA soluble glutamine by HPLC (unit: nanomolePerGram / missing value: NA)
29. ser: five percent PCA soluble serine by HPLC (unit: nanomolePerGram / missing value: NA)
30. arg: five percent PCA soluble arginine by HPLC (unit: nanomolePerGram / missing value: NA)
31. thr: five percent PCA soluble threonine by HPLC (unit: nanomolePerGram / missing value: NA)
32. gly: five percent PCA soluble glycine by HPLC (unit: nanomolePerGram / missing value: NA)
33. arg.thr: five percent PCA soluble arginine+threonine by HPLC (system was unable to separate these two amino acids) (unit: nanomolePerGram / missing value: NA)
34. arg.thr.gly: five percent PCA soluble arginine+threonine+glycine by HPLC (system was unable to separate these three amino acids) (unit: nanomolePerGram / missing value: NA)
35. ala: five percent PCA soluble alanine by HPLC (unit: nanomolePerGram / missing value: NA)
36. pro: five percent PCA soluble proline by HPLC (unit: nanomolePerGram / missing value: NA)
37. gaba: five percent PCA soluble g-aminobutyric acid by HPLC (unit: nanomolePerGram / missing value: NA)
38. val: five percent PCA soluble valine by HPLC (unit: nanomolePerGram / missing value: NA)
39. met: five percent PCA soluble methionine by HPLC (unit: nanomolePerGram / missing value: NA)
40. ile: five percent PCA soluble isoleucine by HPLC (unit: nanomolePerGram / missing value: NA)
41. leu: five percent PCA soluble leucine by HPLC (unit: nanomolePerGram / missing value: NA)
42. trp: five percent PCA soluble tryptophan by HPLC (unit: nanomolePerGram / missing value: NA)
43. phe: five percent PCA soluble phenylalanine by HPLC (unit: nanomolePerGram / missing value: NA)
44. ile.leu: five percent PCA soluble isoluecine+leucine by HPLC (system was unable to separate these two amino acids) (unit: nanomolePerGram / missing value: NA)
45. leu.trp: five percent PCA soluble isoluecine+tryptophan by HPLC (system was unable to separate these two amino acids) (unit: nanomolePerGram / missing value: NA)
46. trp.phe: five percent PCA soluble tryptophan+phenylalanine by HPLC (unit: nanomolePerGram / missing value: NA)
47. cys: five percent PCA soluble cystine by HPLC (unit: nanomolePerGram / missing value: NA)
48. orn: five percent PCA soluble ornithine by HPLC (unit: nanomolePerGram / missing value: NA)
49. lys: five percent PCA soluble lysine by HPLC (unit: nanomolePerGram / missing value: NA)
50. his: five percent PCA soluble histidine by HPLC (unit: nanomolePerGram / missing value: NA)

hf297-04: soil chemistry and metabolites

1. label: label
2. sitename: collection site, study name
3. year: year of sample collection
4. species: tree species
5. subplot: subplot designation within site
6. treatment: treatment
   • HC: hardwood control
   • HH: hardwood high N
   • HL: hardwood low N
7. soiltype: soil horizon
   • Mineral: mineral
   • Organic: organic
8. tissue: sample type and/or analysis
9. soil.ph: soil pH measured in distilled water (unit: number / missing value: NA)
10. per.loi: organic matter measured by loss on ignition (LOI) at 550°C (unit: dimensionless / missing value: NA)
11. per.tn: total nitrogen measured by combustion analysis at 1350°C (unit: dimensionless / missing value: NA)
12. per.tc: total carbon measured by combustion analysis at 1350°C (unit: dimensionless / missing value: NA)
13. ca: exchangeable Ca extracted in ammonium chloride, measured by ICP-OES (unit: milligramPerKilogram / missing value: NA)
14. k: exchangeable K extracted in ammonium chloride, measured by ICP-OES (unit: milligramPerKilogram / missing value: NA)
15. mg: exchangeable Mg extracted in ammonium chloride, measured by ICP-OES (unit: milligramPerKilogram / missing value: NA)
16. p: exchangeable P extracted in ammonium chloride, measured by ICP-OES (unit: milligramPerKilogram / missing value: NA)
17. al: exchangeable Al extracted in ammonium chloride, measured by ICP-OES (unit: milligramPerKilogram / missing value: NA)
18. fe: exchangeable Fe extracted in ammonium chloride, measured by ICP-OES (unit: milligramPerKilogram / missing value: NA)
19. mn: exchangeable Mn extracted in ammonium chloride, measured by ICP-OES (unit: milligramPerKilogram / missing value: NA)
20. na: exchangeable Na extracted in ammonium chloride, measured by ICP-OES (unit: milligramPerKilogram / missing value: NA)
21. zn: exchangeable Zn extracted in ammonium chloride, measured by ICP-OES (unit: milligramPerKilogram / missing value: NA)
22. acidity: exchangeable acidity extracted in potassium chloride, measured by titration (unit: milliequivalent / missing value: NA)
23. cec: effective cation exchange capacity (ECEC) calculated by summation of milliequivalent levels of Ca, K, Mg, Na, and acidity, 100gm (unit: dimensionless / missing value: NA)
24. no3n: nitrate nitrogen extracted in potassium chloride, determined colorimetrically by Ion Analyzer (unit: milligramPerKilogram / missing value: NA)
25. nh4n: ammonium nitrogen extracted in potassium chloride, determined colorimetrically by Ion Analyzer (unit: milligramPerKilogram / missing value: NA)
26. put: five percent PCA soluble putrescine by HPLC (unit: nanomolePerGram / missing value: NA)
27. spd: five percent PCA soluble spermidine by HPLC (unit: nanomolePerGram / missing value: NA)
28. spm: five percent PCA soluble spermine by HPLC (unit: nanomolePerGram / missing value: NA)
29. tube.num: tube number
30. relabeled: new label
31. type: soil horizon
    • Mineral: mineral
    • Organic: organic
32. treatmt: treatment
33. plot: subplot designation within site
34. rep.num: rep number
35. weight: weight (unit: gram / missing value: NA)
36. asp: five percent PCA soluble Aspartic Acid by HPLC (unit: nanomolePerGram / missing value: NA)
37. glu: five percent PCA soluble Glutamic Acid by HPLC (unit: nanomolePerGram / missing value: NA)
38. gln: five percent PCA soluble Glutamine by HPLC (unit: nanomolePerGram / missing value: NA)
39. ser: five percent PCA soluble Serine by HPLC (unit: nanomolePerGram / missing value: NA)
40. arg.thr.gly: five percent PCA soluble Arginine+Threonine+Glycine by HPLC (system was unable to separate these three amino acids) (unit: nanomolePerGram / missing value: NA)
41. thr: five percent PCA soluble Threonine by HPLC (unit: nanomolePerGram / missing value: NA)
42. gly: five percent PCA soluble Glycine by HPLC (unit: nanomolePerGram / missing value: NA)
43. ala: five percent PCA soluble Alanine by HPLC (unit: nanomolePerGram / missing value: NA)
44. pro: five percent PCA soluble Proline by HPLC (unit: nanomolePerGram / missing value: NA)
45. gaba: five percent PCA soluble g-Aminobutyric Acid by HPLC (unit: nanomolePerGram / missing value: NA)
46. val: five percent PCA soluble Valine by HPLC (unit: nanomolePerGram / missing value: NA)
47. met: five percent PCA soluble Methionine by HPLC (unit: nanomolePerGram / missing value: NA)
48. ile: five percent PCA soluble Isoleucine by HPLC (unit: nanomolePerGram / missing value: NA)
49. leu: five percent PCA soluble Leucine by HPLC (unit: nanomolePerGram / missing value: NA)
50. trp: five percent PCA soluble Tryptophan by HPLC (unit: nanomolePerGram / missing value: NA)
51. phe: five percent PCA soluble Phenylalanine by HPLC (unit: nanomolePerGram / missing value: NA)
52. cys: five percent PCA soluble Cystine by HPLC (unit: nanomolePerGram / missing value: NA)
53. orn: five percent PCA soluble Ornithine by HPLC (unit: nanomolePerGram / missing value: NA)
54. lys: five percent PCA soluble Lysine by HPLC (unit: nanomolePerGram / missing value: NA)
55. his: five percent PCA soluble Histidine by HPLC (unit: nanomolePerGram / missing value: NA)

hf297-05: microbial diversity

1. otu.id: OTU identifier
2. h.con.a5.min: hardwood-control-A5-mineral (unit: number / missing value: NA)
3. h.con.a5.org: hardwood-control-A5-organic (unit: number / missing value: NA)
4. h.con.b2.min: hardwood-control-B2-mineral (unit: number / missing value: NA)
5. h.con.b2.org: hardwood-control-B2-organic (unit: number / missing value: NA)
6. h.con.c3.min: hardwood-control-C3-mineral (unit: number / missing value: NA)
7. h.con.c3.org: hardwood-control-C3-organic (unit: number / missing value: NA)
8. h.con.d4.min: hardwood-control-D4-mineral (unit: number / missing value: NA)
9. h.con.d4.org: hardwood-control-D4-organic (unit: number / missing value: NA)
10. h.con.e1.min: hardwood-control-E1-mineral (unit: number / missing value: NA)
11. h.con.e1.org: hardwood-control-E1-organic (unit: number / missing value: NA)
12. h.hn.a4.min: hardwood-HN-A4-mineral (unit: number / missing value: NA)
13. h.hn.a4.org: hardwood-HN-A4-organic (unit: number / missing value: NA)
14. h.hn.b6.min: hardwood-HN-B6-mineral (unit: number / missing value: NA)
15. h.hn.b6.org: hardwood-HN-B6-organic (unit: number / missing value: NA)
16. h.hn.d6.min: hardwood-HN-D6-mineral (unit: number / missing value: NA)
17. h.hn.d6.org: hardwood-HN-D6-organic (unit: number / missing value: NA)
18. h.hn.e2.min: hardwood-HN-E2-mineral (unit: number / missing value: NA)
19. h.hn.e2.org: hardwood-HN-E2-organic (unit: number / missing value: NA)
20. h.hn.e4.min: hardwood-HN-E4-mineral (unit: number / missing value: NA)
21. h.hn.e4.org: hardwood-HN-E4-organic (unit: number / missing value: NA)
22. h.ln.a3.min: hardwood-LN-A3-mineral (unit: number / missing value: NA)
23. h.ln.a3.org: hardwood-LN-A3-organic (unit: number / missing value: NA)
24. h.ln.b3.min: hardwood-LN-B3-mineral (unit: number / missing value: NA)
25. h.ln.b3.org: hardwood-LN-B3-organic (unit: number / missing value: NA)
26. h.ln.c5.min: hardwood-LN-C5-mineral (unit: number / missing value: NA)
27. h.ln.c5.org: hardwood-LN-C5-organic (unit: number / missing value: NA)
28. h.ln.e4.min: hardwood-LN-E4-mineral (unit: number / missing value: NA)
29. h.ln.e4.org: hardwood-LN-E4-organic (unit: number / missing value: NA)
30. h.ln.f2.min: hardwood-LN-F2-mineral (unit: number / missing value: NA)
31. h.ln.f2.org: hardwood-LN-F2-organic (unit: number / missing value: NA)
32. cons.lineage: consensus lineage

hf297-06: OTU rep sequences

• Compression: none
• Format: fasta
• Type: document

hf297-07: gene bank accession numbers

• Compression: none
• Format: text
• Type: document