XYLEM LEVEL README text document

OBJECTIVES
Gather and analyze data about phenotype related to WUE in sagebrush seedlings, specifically, the anatomy of vessels i.e., xylem.

PERSONNEL
Carlos Dave Dumaguit, Boise State University, Boise, ID, USA
carlosdavedumagu@boisestate.edu

Principal Investigator: Sven Buerki, Ph.D., Boise State University, Boise, ID, USA
svenbuerki@boisestate.edu

Stephanie Rude, undergraduate student intern

Walker Morales, undergraduate student intern

FUNDING
US National Science Foundation and Idaho EPSCoR: OIA-1757324

DATA COLLECTION
All raw image data was collected in the Summer of 2021.
Images were manually scored from the Summer of 2021 through Spring of 2022.

After specimen preservation, root cross sections were obtained with xylem cross sections sliced from a section of root within the first inch of the visible soil line on the specimen trunk. 
Dried roots were bathed in water and cross sections were cut and dyed with toluidine blue. 
Cross sections were viewed and imaged in a light microscope. 
Xylem data was gathered by hand as well as no programs existed for distinguishing vascular tissue cross sections for non-model organisms. 
Using the image tool FIJI/ImageJ, diameters of xylem were measured with the vector tool. 
Specific count data was derived from the number of measurements. 
Only one-quarter or 90 degrees of total cross section area was counted to save time.
Polygon outlines of root bundle sheaths were drawn in ImageJ to analyze total vascular cross-sectional area of each seedling. 
Predicted maximal growth of vascular tissue was estimated calculating a theoretical circle of growth using the largest diameter line that could be measured.

STANDARDS
SI - Systeme International (Metric system)

SOFTWARE(S)
R Core Team (2022). R: A language and environment for statistical computing. R Foundation for Statistical
Computing, Vienna, Austria. URL https://www.R-project.org/.
Rasband, W.S., ImageJ, U. S. National Institutes of Health, Bethesda, Maryland, USA, https://imagej.nih.gov/ij/, 1997-2018. 

DATA STRUCTURE
Root folder: 'https://www.northwestknowledge.net/data/92558821-2d5a-4e43-8a96-086c77e564c9/Xylem'
	'./Xylem/00_Raw_data/' - Contains RAW DATA files.
	'./Xylem/00_Raw_data/Secondary/' - This folder contains images of secondary root cross sections. It is structured with folders named after each seedling sample's unique ID.
	'/Xylem./00_Raw_data/Taproot/' - This folder contains images of taproot cross sections. It is structured with folders named after each seedling sample's unique ID.
	'/Xylem./00_Raw_data/TaprootAreas/' - This folder contains images of taproot cross sections for the purposes of area estimation. It is structured with named after each seedling sample's unique ID.
	'/Xylem./00_Raw_data/XylemFunctions.R' - R script containing code that restructures raw images into folders for scoring, creating databases for manual data gathering, and compiles data. 
	'/Xylem./00_Raw_data/DB_xylem_image_metadata.csv' - database that identifies which images are of which seedlings and what type of root cross section was taken by whom.

PROCESSING PROCEDURES/PROTOCOLS
Xylem phenotyping protocol ### need this protocol

KNOWN PROBLEMS/LIMITATIONS

CITATION INSTRUCTIONS
Dumaguit CDC, Buerki S. 2023. Sagebrush greenhouse water use efficiency GxE phenotypic data [dataset]. University of Idaho. https://doi.org/10.7923/jk5q-6876

IP & LICENSING
Creative Commons Attribution 4.0 International ([CC-BY 4.0])(https://creativecommons.org/licenses/by/4.0/legalcode)