Beatriz María Fernández Santos

12 Questions 7 Answers 0 Followers

Questions related from Beatriz María Fernández Santos

NBT 4-Nitro blue tetrazolium chloride, solution...crystals formation overtime?

Hi!! We've been using this product for quite a while now for our whole embryo in situs hybridization with mice embryos: https://www.sigmaaldrich.com/ES/es/product/roche/11383213001 The thing...

31 May 2022 3,826 0 View

Is it possible to CRISPR a plasmid inside a bacteria?

Hi there!! If I have a colony with a plasmid that I've transformed onto it... is it possible to modify my plasmid with CRISPR? So that I can now purify a new modified plasmid? Is this...

23 September 2021 2,108 1 View

Sequencing is not possible after spin column-based nucleic acid purification??

Hi everyone We've been trying to sequence our region of interest from some patients. The genomic DNA they've sent us is not of great quality, but PCR works so they musn't be terrible. I amplify...

30 August 2021 3,416 2 View

How to improve transfection efficacy with lipofectamine 2000 in IMR32 cell line?

Hello everyone I have never cultured cells in my life and everything is new to me. I've done some tests with Lipofectamine® 2000 in my IMR-32 cell line with some plasmids containing dTomato that...

20 August 2021 7,350 7 View

How to make Fast Green FCF from powder?

Hi everyone We need Fast Green to electroporate plasmid into mice embryos, therefore we simply need it to see some color in the liquid we are electroporating so that electroporation is more...

08 February 2021 1,962 2 View

Is there same kind of limit when annealing two DNA products?

Hi there I'm planning on annealing two PCR products to perform some mutagenesis in a plasmid. I was recommended to try and keep it to 600pb each, for example, but for one of the plasmids there...

16 November 2020 8,817 3 View

Has anyone worked with in-fusion cloning?

Hello everyone We've been struggling with our project due to low efficiency rates + check the insert is OK and not in reverse + PolTaq did not introduce a new mutation... and it has come to our...

24 January 2020 6,149 3 View

What should be done differently in a second round of ligation?

Hi, everyone I am using a 4.3kb vector to clone both a promotor and an enhancer. I already have a positive miniprep with no mutations in the promotor, therefore I took some of this plasmid,...

09 August 2019 2,747 3 View

How to improve ligation efficiency?

Hello everyone I've been trying to clone a 1640 pb insert into a 4300pb vector. Transformation works great, after O/N incubation at 37ºC, lots of colonies are found both in the control and in the...

20 July 2019 6,257 7 View

Why does my manual miniprep look like this? What are we doing wrong?

Hello everyone, We are trying to get some constructs after ligation of the plasmid and the insert of interest, and our efficiency rate is very low. Out of 10 colonies, usually only 1 or 2 (or...

16 July 2019 4,011 4 View

Cracked / broken eppendorf after 4ºC 18.000 rpm 10min centrifugation?

Good morning, everyone Yesterday I was performing a manual miniprep. One of the steps consists of a 4ºC max speed (in my case 18.000 rpm) centrifugation for 10 min. When it was done, all of the...

20 February 2019 8,106 1 View

No PCR bands at all?

Hello, everyone I've been struggling for almost a month already when amplifying a promoter from the whole mouse genome: - I already designed 4 primers and have been trying them all (meaning a...

29 November 2018 344 3 View