Juan Pablo Morán

6 Questions 4 Answers 0 Followers

Questions related from Juan Pablo Morán

Can a 5 nt gRNA spacer perform like a 8 nt-cutter restriction enzyme using CRISPR-Cas system?

Or do you know what is the inferior limit in spacer length? 5+3 from the PAM=8

10 November 2021 5,103 2 View

Does site-directed mutagenesis by subcloning works for you?

Hi folks! I want to subclone the S. pyogenes nickase (D10A) gene into certain expression vector by PCR-amplifying it from a Cas9 template and assemble this with a regular isothermal enzymatic...

17 March 2020 4,953 3 View

Do you know what can be the cause for this uniform all-lane smearing?

We are linearizing a vector with a HF polymerase (Q5 from NEB and its recommended parameters). I established conditions that produced a sharp and unique amplicon, but I've just tried it one more...

18 June 2017 4,763 4 View

Do you know the reason for a band in an agarose gel to look like the one at the right in the photograph?Is it a reaction or an electrophoresis issue?

Electrophoresis conditions: 1% agarose, TAE buffer, constant voltage at 7 V/cm electric field strength, and 10 ul were loaded. The amplicon comes from a touch down PCR.

19 May 2017 6,296 4 View

A particular E. coli strain, non deficent in homologous recombination enzymes, should be used for PCR fusion cloning?

We are doing modifications of an expression vector -a promoter substitution-, and I was wondering if attempting this task through PCR cloning using overlaped homologous sequences could be done by...

04 May 2017 7,610 1 View

Does in vivo ligation requieres phosphorylated ends?

We will perform a simple cloning technique based on PCR. We have our covalently closed circular vector and an amplicon produced by asymmetric PCR that will serve as a megaprimer. Hence, it will...

27 December 2016 4,085 5 View