the whole things you did, had to be done, but, there are more important factors that determine the success of your culture. after having the work you mentioned done, you have to follow many other principles during culture and also the treatment you use for explant sterilization is really of importance.... but don't worry some times tissue culture is the story of tedious experiences...  

which plant are you working on? and what kind of contamination are you facing? may you upload a pic of your contaminated culture? 

First; the UV should be for a minimum of 15 min and make sure all items are UV including petri-dishes, forcep (autoclaved), tissue for cleaning (autoclaved), parafilm, filter paper (autoclaved) etc.  

Second; it is necessary to have SPECIAL CULTURE ASEPTIC room, which cannot be entered by anyone and should have specific shoes. 

Third; always use a mask (to cover your mouth and nose).

I guess this will help to reduce the contamination. TQ

Hope u managed all things & you followed right protocols but thing is u didn't wear mask as Mr. Mohammed Sulaimen that thing also affects. Sometimes it happens due to interference of other persons unknowingly.

so don't worry about it try again via following all things sterile equipment's, sterile environment.

Some sources could be pipets used and autoclave not functioning properly. Especially pipets are easily overlooked as source of infections.

a couple of things:

if using agar media in sealed containers, do not open until placing ex-plant and only long enough to place inside. DO NOT open again until you are ready to transfer to another vessel.

If your media contains sugar (for germination or rooting) you will need to transfer within 10 days or when you have 4 axilaries as waiting will allow eventual contamination.

When cloning do NOT add sugar to media as yeast needs sugar and the clones are capable of of making their own sugars, so supply adiquite light. 

also to test flow hood place a sterial container w/ media in flow hood, open for 5 seconds then close and leave in hood to see if hood is functioning properly. 

and everything everyone has suggested already !

good luck 

You should also check the filter and air flow speed of you laminar hood.

Humidity is one of the major factors for contamination. Check humidity level as well.

@Mukund Shukla

Humidity can be. We are controlling humidity by using AC in our culture room. And we are culturing four species in a room. Is this right?

Contamination is a very common thing in tissue culture, especially in hot humid and dusty climate like tropical countries and when using nutrient medium rich in organic substances. In these condition contamination rate might frequently be around 10%. If your rate is higher then you might want to check your equipment, facilities and technique. There are many sources to the contamination and you have to check everything carefully to find out the source.

First of all, check that your equipment, most importantly autoclaves and clean benches are working properly. This can be done by autoclaving some medium bottles, then open half of them inside the running clean bench for about a minute and close. Seal, note them and put all in the growth room for around a month. If contamination occurs only in bottles that were opened in clean bench then it's possible that your clean bench is broken, either air flow is too low or filter is damaged. If bottles that were not opened also have contamination then your autoclave is broken, and doesn't do sterilization of your medium properly.

Secondly, check and clean all your facilities to reduce contamination. Sometimes there are fungi growing inside subculture room, medium storage room or growth room which increase contamination rate. All the rooms should be cleaned, then sterilized with KMnO4 and formol (you don't need 2 kg of KMnO4 and 3.5 liter of formol to sterilize, that's too much lol. Put ~ 1 g KMnO4 in a bottle, then pour ~ 10 - 20 ml formol into the bottle, and run. If your room is big you can make several bottles, or one bottle for each clean bench). During using, try to avoid opening the doors for too long, or entering the rooms while unclean. Lowering the humidity also helps, but usually these facilities are equipped with air conditioners, which already give quite proper humidity.

Thirdly, check that your samples haven't already been contaminated. Sometimes when sterilizing samples from outside into in vitro, the microorganisms are still alive in weaken state and lie dormant on or inside the samples. After 1-2 subculture, they begin to grow out. This mostly happens with bacteria, but sometimes also with fungi. Check your contaminated bottles to see whether in the large majority of them the contamination grows out from where the sample touches the medium. If it is so, this might be the case, and you might want to discard all your samples and introduce new ones.

Finally, improve your technique and protocol and work more carefully to reduce contamination. Turn on UV lamp overnight before work day, sterilize your hand, tools, medium and sample bottles and practically everything with 70% alcohol before you put them inside the clean bench. If the clean bench does not have the front glass to cover your face then you must wear mask to prevent your breath from entering the clean bench. Medium after autoclaved should be put in a clean, dry storage room, preferably near subculture room. Check sample and medium bottles carefully and discard any contaminated bottle before subculture. Contaminated bottles should be gathered and autoclaved before washing, and the autoclave should be cleaned after that. If you are using rubber caps with wool as filter then change all the wool in caps of contaminated bottles. Use sterilized papers to cover the caps or parafilm to seal around the caps to prevent any gap that can let the fungi in. Avoid working during and after rains. Do not go in subculture, medium storage room or growth room after going to places or doing activities with high contamination (gardens, garbage storage, cleaning dusty places, etc.). If even after all this you still have considerable amount of contamination, you might want to try adding fungicides into the medium, or lowering the organic concentration of the medium.

I faced similar issues during my undergrad project work. You can fumigate your working area using copper sulfate crystals and formalin. This would reduce your fungal contamination considerably. You can wipe the working area with a strong disinfectant everyday. Wiping the working area with isopropyl alcohol also helps. Consider keeping the media for sterility check, this can help us to trouble shoot where the source of contamination is. Please extend the time of UV exposure. You can use shoe covers to avoid unnecessary contamination.

May I know what type of yeast contaminant are you experiencing?

You must always start with clean material. We tend to have a lot of contamination from our plant material when we collect from the field at times of the year other than spring. Also, from certain locations in the field. It seems that normally (assuming all of the clean procedures are followed) the source of contamination is from the plant itself. The fungal spores seem to hide in the spaces between leaf primordial and elsewhere. I started washing by rubbing tween 20 directly onto my branches using my fingers to scrub and rinsing before I start my normal bleach cleaning routine. It seems to help.

To avoid yeast contamination hand washing, using sterile equipment, wearing lab coat, head gloves, using sterile laminar hood with 70% ethanol and turning on UV lamp for 10 min and fumigation have been recommended..