Norbert Duró

5 Questions 11 Answers 0 Followers

Questions related from Norbert Duró

Why there is no indel in targeted exons after using CRISPR-Cas9 plasmid PX459? Western blot says there is no protein at the appropriate MW.

I have 5 cell lines now, which has no bands on western blot if I checking my protein. To make sure about the knock-out, I PCR amplified the targeted exons and sent to sequencing (checked the bands...

05 May 2017 2,876 3 View

Annealing oligos for cloning?

I want to anneal two oligos (length: 60bp) to clone it into a vector. They possess sticky ends for ligation. How much oligo should I mix (cc. 100 uM) for annealing?Is NFW suitable or do I need a...

10 October 2016 8,532 2 View

Does my ligated plasmid ready for sequencing and is the purity OK?

I performed a ligation of 1.5 kb insert into a 4.2 kb vector and I got colonies on my kanamycin plate. I picked up 8 colonies and made a miniprep of them. 2 out of the 8 samples contained the...

02 February 2016 2,008 3 View

What could be the problem with the precipitation of DNA with isopropanol during my midipreparation of plasmid?

When elute my plasmid from the column, I get a great amount of plasmid DNA in the elution buffer (~192 ug DNA in 5 ml). After the precipitation with isopropanol (3.5ml) and wash with 70% EtOH (1.5...

12 December 2015 5,604 7 View

lost my pellet after washing with 70% EtOH and centrifuge during midiprep plasmid preparation.

During my plasmid purification with MN midiprep kit I loose my pellet after washing with 70% EtOH and centrifuge at 15,000 5min RT. I clearly see a huge amount of precipitate after using...

11 November 2015 5,482 3 View