surveying polyanthas treated with oryzalin for polyploidy

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david zlesak
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Joined: Tue Nov 26, 2013 12:27 pm

surveying polyanthas treated with oryzalin for polyploidy

Post: # 73215Post david zlesak
Wed Aug 11, 2021 7:13 pm

I love Joe's video tours of his seedlings. He inspired me to record a short video on some polyantha roses I'm working on characterizing for polyploidy. Hopefully you find it interesting. As I was having fun looking at them I had my phone with me and thought it would be fun to share what I was looking at and thinking about. My hope is to try to more easily bridge polys with modern roses. I tried in the past with limited success with more multiflora background polys. These that I treated last year in the video are seedlings that also have a good dose of R. wichurana in them now too. From my limited experience with more wichurana based polys, they seem to bridge and cross more a bit freely (like 'The Fairy') and maybe these tetraploids of wichurana based polys will bridge even better. It'll be fun to find out. Also there are some plants that seem to have 2x and 4x sections and it would be fun to root cuttings and isolate plants of different ploidy levels of the same genotype for additional research in the future (e.g. seeing if there is a ploidy level that is more susceptible or resistant to black spot of the same background like what Andy Roberts in the UK learned with rugosas, etc.).

Here is a link to the video tour: https://share.icloud.com/photos/0vWa2LA ... cAxTFkrsKA

Margit Schowalter
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Location: Alberta, Canada

Re: surveying polyanthas treated with oryzalin for polyploidy

Post: # 73216Post Margit Schowalter
Thu Aug 12, 2021 12:05 pm

Thanks David. Good interesting video.

Jwindha
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Location: SC - Zone 8a

Re: surveying polyanthas treated with oryzalin for polyploidy

Post: # 73218Post Jwindha
Thu Aug 12, 2021 3:42 pm

Very cool!

I attempted at-home, non-laboratory doubling of some practice seedlings using the herbicide Weed Impede as an oryzalin source. I created a 5 uM solution from the herbicide and applied it to the newly opened cotyledons for 12 hours.

For a 1 pint container of Weed Impede with 0.5 lb active ingredient orzyalin:
0.4 ml Weed Impede into 1 L distilled water to yield Dilution 1.
9 ml of Dilution 1 into 1 L distilled water to create second and final dilution, which is the 5 uM solution.

These calculations will be inaccurate if your oryzalin source has a different stock concentration.

I used a graduated cylinder and a handheld engraver to graduate empty liquor bottles. Not the most accurate, but works for at-home use.

Materials:
Weed Impede (active ingredient Oryzalin):
https://www.amazon.com/gp/aw/d/B000I1O7 ... prod_image

Glass pipettes:
https://www.amazon.com/gp/aw/d/B07XM9VL ... prod_image

Plastic graduated cylinder:
https://www.amazon.com/gp/aw/d/B00AITZU ... prod_image

References:
“Chromosome doubling in a Rosa rugosa Thunb. hybrid by exposure of in vitro nodes to oryzalin: the effects of node length, oryzalin concentration and exposure time”
https://pubmed.ncbi.nlm.nih.gov/17641861/

“Oryzalin-induced chromosome doubling in Rosa and its effect on plant morphology and pollen viability”
https://link.springer.com/article/10.10 ... 003-1374-1

“An amphidiploid of Rosa banksiae and Rosa laevigata induced by colchicine”


-Jonathan
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roseseek
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Location: Zone 9b Central California, Sunset Zone 15

Re: surveying polyanthas treated with oryzalin for polyploidy

Post: # 73219Post roseseek
Thu Aug 12, 2021 3:56 pm

Cool! If you need/want 86-3 or any OP seeds of it, please let me know. I've also raised several seedlings using its pollen. All are on HMF. There are two L56-1 X 86-3. One is significantly larger plant with much larger flowers. The other is smaller in all parts, but still a largish climber. The only "repeater" is the Lynnie cross, which I no longer have.
Kim
California Central Coast
USDA Zone 9b
Sunset Zone 15
Cooler inland coastal valley with strong marine influence

mntlover
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Re: surveying polyanthas treated with oryzalin for polyploidy

Post: # 73220Post mntlover
Thu Aug 12, 2021 7:11 pm

Very interesting!
I haven't worked with polys yet. Could you explain some of the difficulty faced in bridging with moderns? Is it similar to sterility issues when crossing moderns with Rugosa? Or do they simply not combine well genetically?
Duane

david zlesak
Posts: 475
Joined: Tue Nov 26, 2013 12:27 pm

Re: surveying polyanthas treated with oryzalin for polyploidy

Post: # 73224Post david zlesak
Fri Aug 13, 2021 10:40 pm

Thanks Jonathan for sharing your well described technique. Duane, I've struggled a lot getting seed set between the multiflora based polys crossed with modern tetraploid shrub roses. They do cross pretty well with Ralph Moore's tetraploid minis though (mainly with his more fertile minis as moms). I have just one successful hybrid from years ago that was generated between a multiflora based polyantha mom (diploid) x 'Champlain'. I'm excited to try these more wichurana based tetraploid polys with modern 4x shrub roses as moms and see what happens hopefully next year. My hope is to root branches from the plants with polyploid characteristics soon and grow them on in the greenhouse this winter where I can count their chromosomes, look at pollen under the microscope, and collect and freeze pollen from those that seem like they may be more fertile. I've used the multiflora based polys with rugosas and got lots of seed set with rugosas as moms, but the seedlings tended to grow weakly and die young. There is one that I saved that has a little fertility and then a seedling of it that has a bit warmer color, so I haven't given up. Growing op polyantha seed from plants growing near rugosas (mainly 'Henry Hudson') I've found some seedlings that seem to be natural hybrids too. I've only used some of the diploid wichurana influenced polys in a limited manner, but was impressed to actually get some Carefree Beauty x Pink Gnome seedlings years back when I couldn't get any seedlings with CB or other easy females with the straight diploid (or even tetraploid) multiflora based polys I mainly had at the time. I'm excited for what may be possible with these new 4x wichurana based polys.

Larry Davis
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Re: surveying polyanthas treated with oryzalin for polyploidy

Post: # 73255Post Larry Davis
Wed Aug 25, 2021 10:11 pm

I like the idea of making your own graduated measuring devices. I found that olive oil bottles make perfect 250 mL graduated bottles. And with a kitchen balance, I don't even need to bring home a graduated cylinder from work or buy one. I weighed out masses of 50, 100, 150,200,250 g on the tare wt of a well cleaned olive oil bottle. I simply used a nursery marker for miniscus marks.

More recently I found that a bottle of graded Kraft or Aldi parmesan cheese has ridges naturally placed correctly. So no marking need if plastic is OK for your work.

Don
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Joined: Wed Dec 31, 1969 7:00 pm

Re: surveying polyanthas treated with oryzalin for polyploidy

Post: # 73259Post Don
Fri Aug 27, 2021 11:59 am

David, thanks for this interesting video and your explanation of what to look for as key characteristics of a plant with doubled chromosomes.

Your efforts have used fresh seedlings. Do you have any advice for technique when trying to double the apical meristems of more mature woody plants like fruit trees? Would timing make a difference with respect to the best season? Would you need more or less concentrated solution of oryzalin, or perhaps the use of DMSO?
What doesn't kill them makes them stronger.

david zlesak
Posts: 475
Joined: Tue Nov 26, 2013 12:27 pm

Re: surveying polyanthas treated with oryzalin for polyploidy

Post: # 73262Post david zlesak
Fri Aug 27, 2021 4:18 pm

Hi Don!

That's a great question. It seems harder with all the leaf primordia over the growing point. Dr, Leo Dionne developed a technique named after him with potatoes years back (1960's I think). He was interested in roses in his later years and was part of RHA for a bit. He would advocate for taking a razor and cutting into an axillary bud so the chemical soaked in, using a saturated cotton swab (with chemicals) for a day or so wrapped around the stem, and then wait to see what grew out. The main growing point was typically killed of the bud, but some to the side buds to the main bud would develop some. There'd be multiple shoots that come out and he would screen those. Years back I tried that with some polys and trifluralin (with DMSO and a drop of soap, etc.). I got stressed buds and those that grew often had multiple shoots. It seemed like only part of some leaves were doubled that expanded and it was hard to find a stable total 4x shoot. Maybe it was hard to get the chemical soaking in as far as needed? Compared to seedlings, there was a lot of power to force out the shoots, which is nice. I'm not sure about increasing DMSO. It really stresses and damages tissues. I think the key could be to focus on getting the chemical into the tissue itself. Maybe a syringe would be better than lots of scraping and cutting with a razor, or combo of both? I like the idea of a moist cotton ball to keep the stressed and wounded tissue from drying out at first so the chemical can soak in and be effective. I'd really love to double 'Caldwell Pink' to try to see if it restores fertility and maybe the partial rose rosette resistance it has can be transferred to the next generation. I suspect for a season, a time of the year with moderate growth and moderate to cooler temps may be best (spring or fall). Active growth is important to have cells in mitosis to interfere with and then at cooler temperatures it slows things down a little bit and lengthens the cell cycle for division giving more time for the chemical to act and interfere with spindle fibers.

Plazbo
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Re: surveying polyanthas treated with oryzalin for polyploidy

Post: # 73276Post Plazbo
Fri Sep 03, 2021 1:05 am

Slight tangent, but did the more multiflora polys that didn't cross well with moderns, stem from the angel rose/wings seed? I know you had selections of those in the past. Probably doesn't help at all and who knows how distant from each other the seed strains are at this point, but may be of interest to someone...they accept r. glauca pollen resulting in seedlings with the same cold greyish top and red under side foliage. Granted no idea if that would assist at all in bridging or fertility at all at this point.


for colour comparison
top left - derived from angel rose/wings seed, top right - some (runty) things from sweet spot calypso op
20210903_172240.jpg

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