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HA tagged transient protein expression

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ranjeet

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Hi,
Friends. I have transfected cells with HA tagged - PtdINs kinase. But the western blot coming nice band at 10 kD above desired mark, that is 55 kDa. But it is single nice band. some times this is coming in control also. Control means nontransfected cells lysate. Could you plase help me out in transient transfection with electroporator and western analysis of expressed protein and plz also give suggestion about how we can increase expression of transfected plasmid. My plasmid in pEGFP N1 substituted with HA in place of GFP.
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Posted Aug 09, 2007, 5:23 AM
saswati1

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What type of tranfection reagent are you using?
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Posted Aug 10, 2007, 0:52 AM
ranjeet

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saswati1 said:
What type of tranfection reagent are you using?


I am using RPMI medium as transfection medium which is without dextrose, b-ME, Antibiotics and without FBS. This has 50 mM Trehalose and 1.25 % DMSO.

Could you please help me.
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Posted Aug 10, 2007, 4:12 AM
ranjeet

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ranjeet said:
saswati1 said:
What type of tranfection reagent are you using?


I am using RPMI medium as transfection medium which is without dextrose, b-ME, Antibiotics and without FBS. This has 50 mM Trehalose and 1.25 % DMSO.

Could you please help me. pH of meium is 7.4 only.
.........................
Posted Aug 10, 2007, 4:13 AM
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