Stage III: Growth of Microgreens in Terrestrial Environments¶

In this stage, participants focus on growth patterns of microgreens in soil growth media. Key data collections centre around growth rates, leaf cover density, and edible biomass. Future space missions will require crews to have access to fresh nutrient-dense crops that are able to be grown quickly, easily, and in limited space. This inquiry seeks to highlight microgreen crops that may be utilized in future space mission studies.

Length of Stage: \~14 Days

Supplies:

For a quick easy protocol you could try Hamama kit, or create your own rooting system.
Standard “1020 Planting Trays” or Standard lab weighing tray, or Petri dishes
Record your experimental design, in your "science journal" or try a online platform such as Benchling.
Two two seed types need to be grown in parallel for comparison
Seed quilts/Planting Soil / vermiculite / perlite / and water (volume is crucial)
Dry Seeds (Any species can be used see supplementary tables for guide on difficulty levels)
Ambient Temperature Thermometer
Digital Scale (sensitive to 0.1g)
Water mister or spray bottle
Camera / smartphone
Label or Tape
Permanent Marker
Ruler & /or a Printable Color (Red) Scale
Computer capable of running Easy Leaf Area Program (if you have a android device you can get instant measurements &/or save photos for analysis on a computer or for sharing with your fellow researchers)
Data Collection Spreadsheet Template (Provided or build your own using our Epicollect5 Class full of researchers program or use our EpiCollect5 AIR MicroGreen Easy Leaf Area)
Microgreen Density Calculator https://jscalc.io/calc/vqHxgaGIt3RncroW

Figure 10: Example of simple microgreen hydroponic and soil media. Source: Photography by Dr Richard Barker and Dr Christina Johnson.

STAGE III Protocol Leaf Analysis ¶

Preparations to be conducted prior to starting: Download and install Easy Leaf Area and Easy Leaf Canopy Free on android devices. Alternatively, plants can be imaged with cameras and then the images transferred to a PC or Mac OS for subsequent analysis.

(See Getting Started on Easy Leaf Area section for details).

Print red color scales (These files are included in the Easy Leaf Program file) this makes it is easier to measure your greens (alternatively you can add the red square manually later but this can introduce errors if the scale is wrong).
Gather trays depending on the amount of growing groups (Ex: Four groups, two trays each), check they don’t leak.
Check light intensity, temperature and any timers or time-lapse photography devices you plan to use.

Figure 11: In this example the microgreen tray has been placed on the button rack under white LED’s. This enabled the researcher to place the camera in front and above to perform time lapse photography. Screenshot of the “Google science journal” in use photographing the microgreens and recording the light intensity and background audio levels.

STAGE III: Microgreen Planting Protocol¶

Use the Hamama kit instructions, or make you own rooting system.
If not using the kit, please precisely record any variations in your protocol using your “Science Journal”.
Fill a single tray with dry soil and weigh (Record the weight of this tray and use it as a reference for subsequent trays).
Pick out dry seeds to be used (At least two different seed types).
Weigh appropriate amount of seeds for the selected tray size.
Utilize the microgreen density calculator to measure ideal seed quantities:
Have participant groups spread their soil evenly in each observational planting tray.
Add water to growing trays. (Soil should be damp, but not flooded)
Evenly spread selected seeds throughout specified trays per seed type. (Seeds can also be spread in rows within the trays. If multiple seeds will be represented in single trays, it is important that each row is properly labelled.)
Gently scattering on the surface of the soil (or growth media). Seeds shouldn’t be covered by soil so don’t plant them deeply. Ideally just a light dusting of soil over the top, put some of the dry soil in a flour sifting sieve and tap the side to get a light dust of soil over the seeds.
Mist planted seeds with water.
Make sure to label each tray with the seed type, date planted, and elevation.
Place near a light source (Windows, or LED lights. Specify light source in datasheet).
Mist soil daily in order to keep it damp.
Measure the light intensity above your plant canopy. An easy way to do this is download the “Google Science Journal Application ''. This tool allows you to use your smart device's camera to measure the light intensity. Try to get the device right in the center of the tray. You can even create a map to show the light distribution over your entire canopy.

(A) Simple 4 tray microgreen hydroponic system under fluorescent lights in the corner of a classroom. This teacher cleverly built it into their skink area to make it easier to wash away the dirt and provide easy access to water.

(B) Along with the soil method, this research group built a simple hydroponic system using an airstone, LED headlight and plastic trays to grow mizuna.

Figure 12: (A) Simple 4 tray microgreen hydroponic system under fluorescent lights in the corner of a classroom. This teacher cleverly built it into their skink area to make it easier to wash away the dirt and provide easy access to water. (B) Along with the soil method, this research group built a simple hydroponic system using an airstone, LED headlight and plastic trays to grow mizuna. This system, while basic, provides valuable data regarding biomass and growth rates for microgreen species.

Software assistance for data harvesting and leaf measuring ¶

Data is to be recorded on the 4th, 8th, and 12th day after planting.
All data is to be recorded in the provided spreadsheet template.

Data Collection Protocol¶

Data Points to be collected:

Top view photograph of tray with printed red scale
Height of plants in cm
Leaf area in cm² (Utilizing EASY LEAF AREA)
General observations (plant health, coloration)
Edible biomass (Weight g)

Photographing Microgreens Protocol:

Print and glue red square scale onto white cardboard or paper
Place red scale next to and at the same height microgreen tray
Photograph the tray at a distance of 45cm from the top of the tray
Upload and label photographs with Plant Name, Date, and Time
Do this on the 4th, 8th, and 12th day after planting

Measuring Height Protocol:

Place ruler at soil level
Using a second ruler (this can also be a pencil, firm cardboard) find the highest point of the plant in the microgreen tray.
Record measurement in cm in the provided data-collecting spreadsheet template
Do this on the 4th, 8th, and 12th day after planting

General Observations Protocols:

The provided general observations template document has sections to record the following measurements:
Ambient temperature at the time of recording
Elevation
Date and Time of observations
In the provided template observations document, participants should record observations regarding all data points collected as well as qualitative records regarding the following points:
Rate of growth
General plant health
Growth distribution within a given growing tray
Plant coloration
Do this on the 4th, 8th, and 12th day after planting

Edible Biomass Measurement Protocol

After all observations have been collected on the 12th day after planting, edible biomass measurements are to be recorded in the general observations document for each tray separately.
Harvest microgreen by removing them from soil media then cutting them at the base of the plant (the point before the roots grow out from)
Each separate tray harvest is to be measured in grams and recorded in the general observations document.

Easy Leaf Area Data Protocol summary

Take photos on the 4th, 8th, and 12th day after plantin
g (or any 3-5 time points)
Follow the EasyLeafArea Protocol to analyze images

Easy Leaf Area Protocol¶

WHAT IS EASY LEAF AREA

Easy leaf area is free, open source, software that rapidly measures leaf area in digital images (photographs or scanner images). Easy leaf area uses the RGB value of each pixel to identify leaf and scale regions in each image

GETTING STARTED:

This program analyzes single jpeg or tiff images or batches of images for leaf area.
Images must have a red scale of known area in the image in the same plane as the leaves for the program to use as a reference scale (This can be added in photoshop or other “Paint” software packages).
Download and install Easy Leaf Area on a designated computer or Android device application.
This software runs on Android as an app, or can be used with images moved to a PC, linux or MacOS.
https://github.com/heaslon/Easy-Leaf-Area
Follow the instructions on their website: https://github.com/heaslon/Easy-Leaf-Area
If you used an AstroBotany Spectral calibration sticker then you should be able to used the new SOAPP image analysis software.

https://github.com/lvbauer/soapp-tutorial/blob/main/soapp-guide.md

Note: more work is needed expanding this part of the tutorial.

Data Analysis Protocol¶

Plant leaf area data

Student Data Entry

Goal: Observe a real experimental data matrix and then make your own

Students are given an example spreadsheet
Students can use these demo data to practice using these analysis tools.
Eventually, the students will enter their own data into their class's shared spreadsheet and then analyse it using these applications.

Numbers are entered into a Google spreadsheet, attached link.

Goal: Students make graphs using transitional tools like spreadsheets and “R-shiny” applications. These tools are specifically designed to process data to produce standardized and flexible graphs.

How to make a Boxplot R-shiny
Play with data in GGPlot R-shiny
How to make a bar plot using R-Plotly
New root growth analysis tool called root RootFit

Statistics

Goal: Introduction to statistics for teachers and students T-test

This article describes how R-shiny statistics app’s can be used in the classroom
How to do a simple T-test using R-shiny app.

How to share Plant leaf area data

Student Data Entry

Goal: Observe a real experimental data matrix and then make your own

Primary data
Environmental meta-data to try and measure.
Temp
Humidity
Light quality
Light intensity
Irrigation system

Numbers entered into a Epicollect5 website or Smart device app attached link.

https://five.epicollect.net/project/urban-microgreens
You can also download the Epicollect5 app onto your smart device and then search for the “Urban microgreens” project.

Data analysis