Pollen systems is simple  and generally the invitro work is carried on on slides using various nytrients ,sugars,boron etc on sildes  which  are maintained at destrable temperatues. The technique is called as inverted drop technique on which the pollen garins grow, Seravh Google for PDFs,Good luck

Dear Sir, I am using surface culture method for in vitro pollen growth. This method allows gradual hydration of pollen. The culture medium (CM) also contains the salts of Ca, B, K, N, Mg which are required for growth. Our problem is not pollen germination  rather the too much coiling of pollen tube which could be due to hardness of culture media. I can't use agar because this solidifying agent is not compatible with PEG-6000 which is used as osmoticum in culture medium. I have also tried agarose. Solidification is achieved only by  Gelrite (0.2%). and tube coiling is prominent in gelrite culture media than CM (without solidifying agent). I can't use CM alone because pollen grains will be more suspended in media and make measurement of tube length difficult

Dear Neha,

did you try the Heslop-Harrison (1979) media which is appropriate for Angiosperms ? It was developed for rye.

I do not work with cereals, but I used it for tree Angiosperm pollen, and it worked well. I looked at your slides, and when I see that the pollen tube do not develop, and some leak, I think that the osmotic pressure is insufficient. For example, for our Betula pollen with Heslop-Harrison media, we use only 0.6% agar, but 200g/l sucrose.

Hope this help.

http://aob.oxfordjournals.org/content/44/suppl_1.toc

Please refer Pollen Biology manual by KR Shivanna .He can be contaccted at the following mail.I  am certain you will get good response.Mention my name:

Prof.Shivanna K R,Asoka Trust,Bangalore

[email protected]

Dear Neha ,

You please try  Brewbaker  and  Kwacks medium.Majority  of the pollen of Angiosperms germinated in that medium .You please refer the text POLLEN BIOLOGY, A LABORATORY MANUAL by Shivanna & Rangaswamy-1993.

I told you earliet and now Dr Sreekala shares. the ssme with more info,

Dear Sir/ Madam, I am already following the "POLLEN BIOLOGY, A LABORATORY MANUAL by Shivanna & Rangaswamy-1993" . Moreover, with brewbaker and kwack medium, the wheat pollen did not germinate. Some cytoplasmic ooze has been seen during pollen growth at 4-5 hour of incubation.

 Dear  Madam Colas, could you please specify the media components (of Heslop- Harrison media) as well as their concentration. This issue of AOB is not available in our library and the article is not an open access article.

Thanks for your help

With Regards

Neha Gupta

Dear Neha,

for rye, the optimum media is :

1% agar, 0.6 M sucrose, 10-3M boric acid (H3BO3), 1 to 2.5 10-3M Ca(NO3)2, 4H2O. pH is not specified, but when you prepare this solution it is around 5.8 to 6.

Fabienne

@ Neha,

Even you have a right medium with right components, you might still not get pollen germination in the correct way, because you added a stress source (PEG6000) and Gelrite in it. There is a possibility that no germination or 'coiling' of 'pollen tubes' or the 'ooz' substance around the pollens might be caused by PEG or Gelrite. Different controls might needed. For example, have you had 'control' plates without adding PEG, and to see whether majority of your wheat pollens germinated and pollen tube grew straight?

Dear Dr Yau, among the media components, PEG concentration has been standardized for wheat (using 20-30 wheat genotypes by a Researcher group). I am following their protocol... I have already tried the media without PEG, pollen germination was not observed in such controls. PEG is actually create an "osmoticum environment" that allow pollen tube growth

@ Neha Gupta

I am facing the same problem as you. have you found a solution for the incompatibility of the PEG6000 and the agar ?