Ethanol precipitation and media preparation blog#3

 Introduction 

This previous week, I helped with an ethanol precipitation and made some media. Ethanol precipitation from what I understand is a process used to “draw out” (precipitate) DNA or in our case RNA. The results were not what we hoped and ultimately had to begin from step one where we needed to innoculate some tgy plates with D. caeni. We made 250 ml of Tgy and LB. 


Methods and results

We started the ethanol precipitation on Monday the 5th and added 150 µl RNA into 6 tubes. Then we proceeded to add 500µl of ethanol, along with 20µl sodium acetate. The last thing we added was 2 µl of glycogen blue to be able to see the pellets the next day. We left the tubes in -80 deep freeze overnight. Then on Tuesday, we put the tubes in the centrifuge at full speed for 15 minutes. While this process took place we made 250 ml of Tgy and LB. we measured out the ingredients


Tgy LB

-Deionized water 250mL               -Deionized water 250mL

-yeast extraction .75g                   -Tryptone  2.50g 

-Dextrose .25g                              -Yeast extraction 1.25

-Tryptone .75g                               -NACI 2.50g

-Agar 3.75g                                   -Agar 3.75g 



While I was making media, Madison, proceeded to take out the tubes of the centrifuge and wash the pellets then spun the tubes for 10 minutes, and did processes twice. Then we let the tubes air dry in the fume hood, madison then added RNase to each tube and vortex each. Then we measured the concentration of the tube with the nanodrop and did not get The result we hoped for. 


Discussion 

I'm not really sure where we went wrong and why we weren't able to get the results that we wanted. Maybe we did a step wrong in the ethanol precipitation process but I don't even know which one. We decided that we would have to start the whole experiment again from inoculating some plates with D. caeni. We were supposed to do a DNA isolation later on in the week but I was not feeling well and was unable to attend the lab on Thursd

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