PhytoAgar - International Seed Testing Association · • Eliminates blotter/paper media toxicity issues • Can be used in current germ testing containers • Vacuum planters and

PhytoAgar The Next Generation Germination Media

Presented by David M. Johnston, RST - Monsanto Veg. Seed Phys. Lab

v01MAY2014

AGAR Germination Media of the FUTURE!!!

(….it might be closer than you think???)

Brief Overview • Agar is extracted and processed from seaweed, which is a

“plant-form” of specific red algae species.

• Discovered in 1600s by Minoya Tarozaemon, a Japanese innkeeper, who was said to have discarded some surplus seaweed soup and noticed that it gelled later after being discarded.

• Today various types of agar are commonly used for culinary and scientific purposes (e.g. microbiology, tissue culture, etc).

• Agar is already successfully used for seed germination at Kew Gardens in the UK and by other labs in the US for vigor tests.

• Next application will be for mainstream seed germination testing.

Agar Prep – Simple as Boiling Water •Heat water to approx. 100˚C using microwave, hot plate, autoclave, etc. (Agar melting temp 95˚C/203˚F )

•Add appropriate grams of agar powder for desired concentration and stir until fully dissolved. •Gently cool agar solution to approx. 50˚C/122˚F or lower for safe handling. (Agar gelling temp 38˚C/100˚F)

Agar – Pouring •Agar solution will start to gel at 38˚C/100˚F so the temp must be monitored as the solution cools. •Agar solution must be promptly poured into containers before gelling starts. •If agar gels prematurely reheat to liquefy. •Solution should be poured carefully as to not create bubbles in the agar. •Work surface must be level to allow agar to obtain an even depth in the container. •Once agar has solidified (approx. 10 minutes depending on thickness) it should be covered. •Incidental condensation may form inside containers but does not require mitigation. •Once agar has gelled and cooled it is ready to use or may be be stored in a cool chamber a short time for future use.

Pouring agar in the lid.

Pouring agar in the box.

Agar – Planting

•Standard vacuum planting options and counter boards are suitable for use. •No special equipment required for planting. •Standard germination containers usable. •No special media cutting/ordering as with paper based media scheme.

•Agar is ALWAYS the right size!!!

•No media toxicity issues!!! •Numerous agar vendors available. Works great on lids

even when they are not uniformly flat.

Agar – Planting

Pepper Tomato

Lettuce Cucumber

Agar – Behind Closed Doors

There is NO “media shadow” with agar as there is with paper!!! Light exposure is maximized with agar since it is a clear media.

Agar – Seedling Development

Top view 4inX6in Box

Bottom view 4inX6in Box

Pelleted tomato

(Equivalent to 10cmX15cm)




Tomato





Brassica sp.





Lettuce





Cucumber


Agar – Seedling Evaluation

Poor seedling remove technique during the first count may result in agar disruption at first count.

Pelleted tomato


Proper seedling remove technique will result in minimal agar disruption at first count.

1

2

3

Pelleted tomato


Even with poor seedling remove technique during the first count and resulting agar disruption, the seedlings still developed normally.

Pelleted tomato


Other possible options to reduce media disruption at first count. •Formulate a denser agar so roots do not penetrate the media during germination for easier seedling removal at first count. •Slant containers vertically to cause roots to remain on top of agar and orient vertically for easier evaluation and seedling removal.

Pepper

Agar – Clean Out

Agar clean out is very simple and easy to accomplish since agar does not adhere to the container, unlike organic media and sand.

Instructions: •Tilt container and insert a thin flat utensil under agar layer in the corner of container •Gently push utensil forward lifting and pushing agar out of container into waste receptacle •Wash container as usual after use

Agar – Let’s Do The Math Common recipe:

10 gr agar powder/1000 mls water (1% conc.)

Usage:

• 4X4 box lid - 50 mls agar (Agar cost $0.05 per 4X4 box - Based on the 5Kg price of $504.30 USD with 50 mls

of 1% agar concentration used per box.)

• 4X6 box lid - 100 mls agar

Example cost of agar powder: Suppliers:

(These are all online prices APR 2014)

Agar – Pros/Cons • Labs of various sizes are successfully using agar in USA and

other world regions (Kew Gardens in UK) • ISTA currently considering validation as a media • Very homogeneous and consistent non-toxic product

facilitating greater testing uniformity in and between seed labs

• Eliminates blotter/paper media toxicity issues • Can be used in current germ testing containers • Vacuum planters and counter boards can be used • Evaporation rate less than paper media and sand • Reduce/eliminate need to add water at intermediate counts • Since prepared agar absorbs water, thus additional water

may be added if needed during test period • Provides more even water absorption by seeds, potentially

reducing imbibition injury • Light exposure is maximized in germinators since agar is a

transparent media creating no “media shadows” • Seedling roots are fully visible even when roots penetrate

agar, unlike paper media • Agar is inert and does not encourage or support pathogen

growth • Media additives (e.g. GA3, KNO3, etc.) are easily

incorporated during agar prep • Significantly reduction of lab waste as compared to paper

media • Very easy to prepare agar with very minimal basic

equipment requirements – heat source for water required and thermometer

• Seedlings exhibiting primary infection can be easily removed from replicates during test period

• No special media sizes required to be cut or ordered and kept in stock by labs

• May not be appropriate for all seed kinds • May not be easily accommodated by all lab testing

schemes • Safety precautions must be followed with handling

hot liquids • May be a increase in time for media prep • May require increased planning time for media prep

due to agar prep required • May need to purchase equipment for heating water

for agar and appropriate thermometer • May require increased media prep area

Agar – Let’s Talk About it

• More research is needed to validate. • May not be appropriate for all crops. • What agar concentration works best? • How think should the agar bed be for various seed sizes? • Can agar be reheated and reused? • Agar is already being used in seed labs. • What is your current view with using agar for testing? • What are your likes and dislikes? • What are your concerns? • What are your unanswered questions? • What is your advice on how to proceed? • If research across several labs validate agar as a germination

media, would you support it being added to the ISTA Rules?

THE END

Documents

PhytoAgar - International Seed Testing Association · • Eliminates blotter/paper media toxicity issues • Can be used in current germ testing containers • Vacuum planters and