Does Puromycin work without polyA signal after it? - (Aug/12/2014 )

Hi All,

I am new to the mammalian expression system. Recently, I was working to make specific point mutation on the mouse genome with CRISPR. I have made a HDR donor construct "gene 5' Arm----PGK---Puro----3' Arm" but without Poly A after Puro. My question is whether I can use puromycin to screen my ESCs. I am worried about Puro does't work without PolyA. Thank you so much in advance. 

Zero

zero85 on Wed Aug 13 04:17:33 2014 said:

Hi All,

 

I am new to the mammalian expression system. Recently, I was working to make specific point mutation on the mouse genome with CRISPR. I have made a HDR donor construct "gene 5' Arm----PGK---Puro----3' Arm" but without Poly A after Puro. My question is whether I can use puromycin to screen my ESCs. I am worried about Puro does't work without PolyA. Thank you so much in advance. 

 

Zero

Could you give the whole sequence of the puromycin resistance gene you're using? I will check it against what I'm using because I know mine works.