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Does Puromycin work without polyA signal after it?


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#1 zero85

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Posted 12 August 2014 - 08:17 PM

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



#2 seanspotatobusiness

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Posted 04 September 2014 - 05:48 PM

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.






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