While reading Henrys post on microscope calibration it occurred to me (thanks to Ann and David’s comments on screening mesh and sieving pollen) that we could get more predictable ploidy levels in seedlings when using triploid parents by screening the pollen before applying it to the seed parent.
If diploid pollen on the whole is no larger than 40 microns in diameter then using the triploid pollen that passes through a 40 micron mesh would ensure that you are applying only diploid pollen.
Likewise if you desire tetraploid seedlings you could use the pollen that would not pass through the mesh and increase the chances of getting tetraploid seedlings.
If using fresh pollen what would be a safe cut off size if sieving for diploid pollen, and a safe minimum size mesh to hold the tetraploid pollen?
Thanks,
Jinks.
Hi Jinks,
Yes, that is a great idea. Here are a couple references that use mesh to separate pollen. I think though that 40 microns is too large. At least with acetocarmine staining the average diameter for diploid rose pollen is about 32 microns and for tetraploids I found it to be about 40. Perhaps we should measure pollen size in sugar/boric acid water solution used to germinate pollen “in vitro” because that may be a good solution to use for screening and then applying it to stigmas. Maybe water would be fine too. I suppose after screening pollen in solution like that one would want to use it right away. Using dry pollen would probably be problematic because it would be shaped like ovals or footballs and the distinction and seiving ability would be compromized.
Sincerely,
David
Eijlander R (1988) Manipulation of the 2n-gametes frequencies in Solanum pollen. Euphytica 39S:45-50
Okazaki K., Kurimoto K, Miyajima I, Enami A, Mizuochi H, Matsumoto Y, Ohya H (2005) Induction of 2n pollen in tulips by arresting the meiotic process with nitrous oxide gas. Euphytica 143:103-114
Eijlander in the 1988 paper successfully used both water and a “pollination germination” water solution to force the smaller pollen through the screens that he used. Surprisingly (to me at least), the resulting wet pollen could still be used to “pollinate” the flowers.
Since non polar organic solvents are used to store pollen. it would appear, to me, to be better to use an easily evaporated non polar organic as the forcing liquid.
See the following thread for a discussion of the use on non polar organic solvents:
http://www.rosehybridizers.org/forum/message.php?topid=1757#1777
The next question, in my mind, is how to obtain an inexpensive filter of the correct pore size to separate dipold from tetraploid pollen (and not be degraded by organic solvents). Please note the word “inexpensive”.
One possible solution is to use gasoline filters. The following link gives the results of a Google search for “35 micron” filters:
http://www.google.com/products?q="35+micron"+filter&btnG=Search+Products&hl=en&show=dd&checkout=1
Any other suggestion for filters?
So far I am thinking of using a large syring (found in farm type stores) to force a pollen/non polar organic solvent “slush” through a gasoline filter. After the initial liquid (and much of the small pollen) is forced through the filter, one or more addition "rinses with pure solvent could be passed through the filter with the syringe to further enhance the separation.
Link: www.google.com/products?q=%2235+micron%22+filter&btnG=Search+Products&hl=en&show=dd&checkout=1
H. Kuska question: Why not just pollinate with the mixture of 1n and 2n pollen, and then separate the resulting plants by ploidy level?
The following paper suggests that you may get plants pollinated with 1n pollen.
In a study of naturally occuring 2n pollen in white poplar Z. Zhang, X. Kang, P. Zhang, Y. Li, and J. Wang
( http://www.springerlink.com/content/r78v316562g25385/ )
reported: “In vivo germination in stigmata revealed that 2n pollen germinated later and that pollen tubes grew slower than n pollen (Kang and Zhu 1997). This finding suggests that the fertilizing ability of 2n pollen is weaker than that of n pollen and that it has less of a chance of participating in fertilization.”
My (H. Kuska) comment. Thus, is appears that the use of a pollen size filtering / concentrating system would increase the success rate for obtaining 2n fertilization in systems with mainly 1n pollen.
Link: www.springerlink.com/content/r78v316562g25385/