A simple method of embryo culture for plant breeders

In the latest edition of the newsletter I provided a link to download a .pdf version of a manual of technique for the extraction and in-vitro germination of mature plant embryos that I have written. For those folks who are not subscribers (tsk tsk) the url is:

http://holeman.org/embryoculture.pdf

Also in the newsletter I provided a link to download a supplemental chart to accompany my article “Fun with numbers: the species ancestors of modern roses”. The chart is not yet available by that link, but it can be downloaded temporarily at

http://holeman.org/chart.pdf

While the appearance of the .pdf version of the chart on-screen is poor, it will print properly. View the chart as a .jpg on screen at

Link: holeman.org/images/chart.jpg

This is an amazing resource Don. Thank you!!! You have navigated and presented ways to help us extract embryos and take care of them so they can survive through much trial and error. What an amazing tool to be able to help get fast germination of precious seeds and acheive germination from some that we just can’t get to germinate well because of very thick, hard outer coverings or whatever reason is holding them back.

This is an amazing contribution Don and thank you again!

David

Hi Don. Thanks for the article, it has been interesting to read.

Thank you so much Don. I will be having some seeds that I will need to use this on but I will practice on op seeds first.

Patrick

It was a great read, although I’m still reluctant about doing it since I’m not good at calculating solution strengths.

Have you’ve thought about creating a video and uploading it on youtube?

The youtube’s How-To’s section are amazingly entertaining and educational.

Yes, it is a wonderful article. I confess that I’m lazy enough just to sow the seed and wait to see what happens.

This really might come in handy for utilizing hard to germinate seed some time in the future.

Great article! Your method should provide some great entertainment of cold and wet Oregon evenings.

I confess that I’m lazy enough just to sow the seed and wait to see what happens<<<<<<

Robert,

I have directly planted both seed and bare embryo in potting mix…Both naked embryo and naked seed do germinate this way, sometimes the seed will germinate as fast as the embryo.

BUT, you are looking at definfitely less germination percentages overall, if you by-pass the culture step.

In the end it is up to everyone to decide how badly they want high germination rates.

“In the end it is up to everyone to decide how badly they want high germination rates.”

Thanks George, yes, I realize this. I am already so overwhelmed with seedlings, it might be just as well many of my crosses don’t germinate.

As I’ve mentioned before, I often change directions after I’ve made my crosses for the year. I’ll have to think hard to choose amongst those crosses I’ll actually sow this season.

Wonderful presentation Don, I especially like your handy dandy seed splitter! Pretty nifty tool. You really emphazise cleanliness and I believe that is so important in handling your seeds and even more so with the embryos.

They are so delicate. I have not tried your method yet but I will. However I am having white mold form around my embryos, could it be from the water I am using. I use purified bottled water and H202 3% to soak them in before I put them in the baby food jars that George has suggested using. Alot of the achenes that I have opened have had dead embryos in them which is very discouraging but I am glad I did not waste my time with the stratifying for 2-3 months and then the time waiting for them to germinate.

Great work, Don! I’m floored! Will have to try this on the crosses I’ve done with Persian Yellow.

I have been practicing your technique on some open pollinated shrub rose hips. I am extremely good at destroying the things but I did manage to mess up a few times and know have a few embryos under culture. We will see how it goes. I got more of these seeds to practice on. Maybe I will get it down. It is definitely not easy to get the knack of. Thanks Don for the information on this technique. Also thanks for the article on the ancestry of modern roses I really enjoyed it.

Thanks to everyone for all your kind remarks.

Have you’ve thought about creating a video and uploading it on youtube?

Yes, but TV always makes my hands look fat! Actually, Enrique, it crossed my mind but my operation is very low-budget and I simply don’t have a video camera.

However I am having white mold form around my embryos, could it be from the water I am using

Jeanie, this white fungus is more likely to be from the seeds themselves. Usually it is isolated to a particular cultivar so make sure to be meticulous when cleaning between batches of seeds. I’ve had success in dosing young seedlings with Bayer fungicide dropwise at the standard application rate though you do have to be persistant and sometimes you really don’t get rid of it until your seedlings get outdoors into the sunlight and a regular spray routine [donning my tin hat and ducking quickly to dodge the bricks from the Earth Kind breeders].

I am already so overwhelmed with seedlings, it might be just as well many of my crosses don’t germinate.

Robert, given your penchant for species crosses do any of these reluctant germinators hold promise of opening up new avenues for future hybridizers like your work with bracteata?

Don said,

“Robert, given your penchant for species crosses do any of these reluctant germinators hold promise of opening up new avenues for future hybridizers like your work with bracteata?”

Actually Don, I’m passed that point, at least at the tetraploid level. My seedlings seem to be fully fertile.

I’ve got the opposite problem, too many seedlings and not enough room to grow them all out.

These techniques could come in very handy for the future. I can think of a great many situations where they could used to make breakthroughs that might otherwise be impossible.

In terms of producing something commercial looking it will always be a numbers game. We need to grow large numbers of seedlings in order to select for the characters most people desire.

There are still plenty of surplus hips in the garden. I will try my hands on this. Thanks for the information.

I just can’t open those darn seeds.

I’ve tried several methods…

The first time, I tried to chip off the coat. I exposed it down to a fibrous layer, but… I can’t remove it without damaging the embryo.

I’ve applied the blades of the clippers directly on the suitors of the seed length wise… I can remove half the seed in that way, but the embryo is still stuck on the other half.

I’m thinking about using a nut cracker and trying to apply pressure on the ends of the seed and, hopefully, have a clean removal of the “shells.”

I’ve been following Don’s method for some time and have acquired a tubing clamp to do it. I find that if I do the achene straight from the hip I usually squash them (BTW Don… I have been working on a design for seed splitter that you can readily buy from a hardware store… when I get off my bum to buy one and modify it slightly I’ll send you pics once I’ve trialed it to make sure it works). If I let them dry out a few days then they crack open better. I have noticed that even when you don’t follow the aseptic protocol that the embryos continue to develop normally though I may be just lucky because the water to my house goes through a UV steriliser first so is pretty clean form the outset.

I figure this method should not be used routinely, but in special cases when we just can’t get seeds to germinate normally, when we are approaching the end of the season, or when we have small numbers (once one is good at it… I still destroy a lot of them). I think, if possible, it is better for the seed to germinate naturally… a bit of survival of the fittest… I’m using it at the moment on some ‘Altissimo’ seeds because I just can’t get them to germinate. The seeds are so large with such thick walls that I think this is preventing germination… maybe the embryo’s are running out of steam before being able to effectively emerge from the achene or sufficient moisture is not getting into to initiate germination, or they just need a whole lot more time to break down enough to split apart??? This is one I did on Thursday night. The radicle is just beginning to emerge now… so viability of the embryo must be ok… These ones, in the same bag as some Leucospermum seeds, and Serruria seeds, did not have the peroxide dip.

I’ve applied the blades of the clippers directly on the suitors of the seed length wise… I can remove half the seed in that way, but the embryo is still stuck on the other half.

Don’t give up, Enrique. It takes practice. When you get this far a few more snips usually are enough to let you lift the embryo out. Use the side of your tweezers or needle probe to push against the embryo and lift it out.

I have been working on a design for seed splitter that you can readily buy from a hardware store

Great! I found that subtle tweaks in the design made a lot of difference so play around with your concept if at first you don’t succeed.

I have noticed that even when you don’t follow the aseptic protocol that the embryos continue to develop normally

Rose pathogens are more likely to be present in the seeds than in your water supply. The cleanliness protocol I published is a little bit rigorous to employ but it prevents cross-contamination of embryos as much as is it kills the bugs in the water. For me, the higher yields are worth the extra effort.

The seeds are so large with such thick walls that I think this is preventing germination… maybe the embryo’s are running out of steam

Simon, I had similar thoughts about ‘Prairie Peace’ seeds. The seed wall is at least 1/16th inch thick. No wonder Robert Erskine was never able get them to germinate. Robert tried to cross ‘Prairie Peace’ with ‘Peace’ rose but never got hips to set although crossed both ways. Given that Don has developed this method to germinate the seeds, I hope someone will be successful with the cross. It would be a great way to honour Robert Erkine.

Link: www.helpmefind.com/plant/l.php?l=21.87165

Whenever I see seed coats like this I always assume a long stratification period would be required in order to break down the seed coat.

For awhile I was reading a lot about use of Bromelain to break down seed coats. Can anyone offer feedback as to how this worked out?

Also, Margit, Do you know of a source for ‘Prairie Peace’?

The link at HMF for the source in Canada seems to be inactive.

Thanks, Robert