Monika Gostic

21 Questions 21 Answers 0 Followers

Questions related from Monika Gostic

In situ hybridization protocol for SKIN cells (zebrafish)?

I have extensive experience in WISH and FISH in zebrafish embryos when it comes to mapping the expression of genes within the fish. I am now about to start a new project, checking the expression...

04 March 2020 9,947 1 View

Zebrafish Tol2 to test human promoters?

I have conducted a study using Tol2 Gateway system to study human promoter in zebrafish. As per literature, the system should have worked, but it didn't. The zebrafish positive control was...

02 March 2018 2,953 2 View

What would be the best marker to label ALL (or as many as possible) glia cells in zebrafish?

I am identifying cells that express my gene of interest and I think they might be glia cells. I would therefore need a reliable marker that would label as many glia as possible - Muller,...

26 June 2017 5,653 1 View

Is there a zebrafish marker for eye develoment?

I am doing an in situ hybridization and have localised my expression to the developing zebrafish eye. I would like to have some kind of positive control, but am not too familiar with eye...

07 June 2017 282 5 View

Fixing whole mount transgenic zebrafish for Imaging analysis?

I'm doing a large scale analysis of expression patterns in transgenic zebrafish and I am chasing after certain developmental stages. Therefore I need to fix the fish at that stage and once the...

01 March 2017 8,957 4 View

Maximum in situ hybridization temperature?

I am performing a whole mount in situ hybridization on zebrafish embryos. I have increased hybridization T to 65C but am still getting nonspecific binding. Can I increase the T even higher and...

18 January 2017 7,668 9 View

Zebrafish mounting for confocal microscopy?

I am trying to image my zebrafish embryos from 0-120hpf and am struggling with suitable mounting without damaging the specimen. Does anyone have a good protocol or own experience for it? I am...

01 December 2016 3,730 4 View

Gateway Tol2 zebrafish transposone system with human promoter?

I'm using Tol2 Gateway system to test for human promoter activity in zebrafish. I am already using zebrafish UBI promoter as a positive control, but I was wondering if anyone might have a HUMAN...

24 October 2016 2,004 2 View

Long range PCR optimisation?

We're performing a long range PCR, longer than 3kb and so far it has failed to work. We've tried DMSO, Mg2+, touchdown approach, different DNA, new primers, different polymerases. I'd appreciate...

03 August 2016 7,589 8 View

Up-to-date CRISPR knock-out protocol in zebrafish?

Is anyone working with CRISPR/Cas9 in zebrafish and has a detailed up-to-date protocol with high efficiency? I would like to create a knock out of my protein of interest, but so far none of my...

03 August 2016 9,206 6 View

Evaluating Tol2 F0 results in zebrafish?

I am performing transgenesis in zebafish, using a Tol2 system. I have successfully constructed the entry plasmid and the integration of my wished construct has also been confirmed. However, I do...

29 April 2016 5,877 8 View

Can anyone suggest a good zebrafish anti-gamma-tubulin Antibody?

I'd like to visualize zebrafish cilia, for which I'd need an antibody such as the one against γ-tubulin. Does anyone know any good ones? Or if you might have other suggestions how to stain for...

12 June 2015 5,225 2 View

What do I do when my RNA purity is over 3?

My CAP and PolyA RNA has been quantified on the nanodrop and all the samples show high concentration as well as the ration between 260/280. A is over 3 in all of them. Does that mean I have extra...

27 March 2015 6,283 13 View

Does anyone have a plasmid for Otx5 ISH probe for zebrafish?

I'm performing an In situ Hybridization on zebrafish and would need a probe for zebrafish otx5, expressed in pineal and parapineal. Does anyone have it (or maybe something similar). Many thanks!

05 March 2015 5,777 2 View

What do you use as a positive control when performing Tol2 experiment?

I've created a construct for Tol2 Gateway enhancer assay and am about to proceed with experiments. However, I do not have a positive control (nor negative). Can you suggest me what would be the...

26 January 2015 7,180 6 View

How do I obtain a high concentration of my final antisense probe for in situ hybridization?

I created sense and antisense probe by extracting and purifying RNA, generating cDNA, performing PCR for wished construct, which I then cloned into pCR-Blunt II - TOPO vector. After isolating and...

08 January 2015 1,951 2 View

How can you quantify CAP-poly-A RNA?

I have created capped, polyadenylated RNA in order to microinject it into zebrafish embryos. I would now like to make sure that what I've got is the right construct. How do I quantify the...

12 December 2014 9,462 2 View

How do I see how the change in amino acid changes the function of the protein?

I have a sequence of my gene and protein of interest and I have a database of mutants telling me where the mutation in the sequence is. I want to know how this mutation affects the protein...

07 November 2014 4,070 13 View

What is the best positive control for in situ Hybridization of early developmental genes in Zebrafish

I'm performing an in situ Hybridization protocol and have no knowledge of the expression of my candidate gene. Firstly, I will perform a qPCR in order to see when the gene of interest is...

02 October 2014 9,372 7 View

How do you insert a PCR fragment in reverse direction into the vector?

I have a PCR fragment with blunt edges that I'd like to insert into my vector. I already inserted it in a normal forward direction, but now I'd like to try it also in the reverse direction. Does...

25 August 2014 604 5 View

What happens if you mutate Arg into Trp?

I've taken over a project from a previous postdoc and have discovered a mutation on one of the sequences. This mutation is located right after ATG site and it mutates GCC into TCC, causing...

01 January 1970 9,938 0 View