primer resuspension, buffers -step by step
Could you please help me how to prepare TAE (50X) and TE buffer?
I have already found a recipe for TAE here among the threads, so is it the right one?:
"First you need to know preparation of 0.5M EDTA solution:
Add 186.1 g EDTA (disodium, dihydrate, ) to 800 ml of ddH20.
Add about 20g of NaOH pellets while stirring to bring the pH to 8.0.
Add the last few grams slowly to avoid overshooting the pH.
Note that the EDTA won't completely dissolve until the pH is around 8,
make the final volume to 1000ml with ddH2o.
Filter with 0.5 micron filter and autoclave
50 X TAE Buffer Preparation protocol (Tris-Acetate-EDTA)
242 gm - Tris base
57.1 mL - Acetic Acid
100mL - 0.5 M EDTA (shake vigorously before use)
Add ddH2O to 1 Liter and adjust ph to 8.5 using KOH."
I would also like to resuspend our oligos (primers) in TE according to the newletter of the company for long term storage I am to use : TE buffer (10mM Tris, 0,1mM EDTA pH 8.0.)
Since I was given the task to start the molbio lab without an experienced laboratory technician and without lab practice (just with theory backgorund: molecular biologist, got the diploma 10 years ago, and hs worked on different field until now) I need to spend endless time to try to clarify all small questions, and after spending days with reading I am more and more scared....
Could you pelase help me?
Thank you very much in advance,
Re: primer resuspension, buffers -step by step
The 50x TAE buffer protocol is correct. EDTA can only dissolve when the pH reaches 8. Yes, for long term primer stock storage at -20C, it is better to dissolve the lyopholized primers in TE buffer. Then you can dilute you primers in sterile water for PCR work from the stock.
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