If 20000 ppm calcium hypochlorite is not work then you can try physical methods especially high pressure (30-200 Mpa), it might work.

Thanks prosanta

Dear Alsaberi

There may be so many reasons for this issue.If the contamination is appearing on non-inoculated plates, the fungi are coming from improperly sterilized media and/or equipment. If the contamination is appearing at the points of inoculation (on plates) then the fungi are either in the inoculum or on the instruments that you are inoculating with. If the contamination is appearing on the surface of the plates, but away from the places that you inoculated the bacteria with, fungal spores are probably floating in from the air

The most important things you can do to prevent air contamination is to not open the container (either petri dish or flask of liquid medium) in the open air of the lab. ONLY open them under a clean laminar air flow hood that has been running for at least 15 minutes before using. AND when you are working under the hood do not have anyone walk behind you. This can disrupt the air curtains of the hood and throw contaminated air over your work. This last point cannot be ignored. changing the traffic pattern within the room and the problem was eliminated.sometimes the filters applied in laminar flows are overused and therefore the filters are filled with airborne fungi such as those You have implied in your text. The when the pump acts it blows the spores into the chamber and You get contamination as the result. Then check if the filters are enough efficient to be used. This is the most important point I think .You are better to check as You have used sterilized autoclaved/ microfiltered media, then your media shall be sterilized. Another point is the necessity of a UV lamp to keep the chamber sterile, and or at least a flame source. One other step is to add some fungicidal compound to the inoculum aliquot You use to inoculate the media. But still I emphasize on the lifetime of filters of laminar flow to be checked and changed with new ones.

Good Luck.

Thanks Kulvir

Great explanation with good details,

I am testing also other plants such as,Tomato, Corn,Soybean,and Arabedopsis so I don't have any contamination. The only plant that I am struggling with is Sorghum

I think the reason is that the fungi are coming from inside seeds themselves (endophtic fungi). The contamination begins when the sorghum seeds start to germinat and from the seed place only in the plate.

Off course, I think you found the exact reason.Sorghum mostly gets infested when grains reach close to maturity.

I assumed that, but I do not know yet the solution for this problem(contamination)