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Questions related from Kunal Chopra
I genotype and sequnce zebrafish using a bit of the fin. Lately though, I have been struggling with PCR. So far I have tried the following but to no avail: 1. Tried the primer pair in question...
08 August 2018 7,761 5 View
Following CRISPR mutagenesis and sequencing analysis of the F1s, I have found a heterozygous male with 10bp deleted on exon1 and a heterozygous female with 5bp deleted on exon 1. I was wondering...
03 March 2017 3,802 8 View
I have a mutant strain of zebrafish wherein the mutation is considered to be directly affecting hormonogenesis. My fish do show some phenotypic aberrations due to this but otherwise seem healthy...
01 January 2017 8,018 8 View
I need to do an ELISA assay of a hormone on some tissue samples from zebrafish. Existing literature suggests homogenising tissues in ice cold phosphate buffer. However, I found out that PBS,...
09 September 2016 3,399 4 View
I am testing zebrafish and Xenopus laevis for reduced hormone levels. It is the same hormone in both species. My method of choice is ELISA. However, when I researched for the available kits, they...
08 August 2016 4,551 3 View
I have a mutant zebrafish strain that likely has hypothryoidism as a result of the mutation. I am basing this on the fact that the adults are dwarfs compared to wild type fish and also do not...
05 May 2016 7,426 2 View
I just did a total RNA extraction using TRIZOL and the 260/230 values of all samples is between 0.50 and 0.90! I also noticed that for some of them the pellet did not dissolve completely. Also, I...
04 April 2016 5,655 7 View
During PCR, is there a specific parameter which helps determine the temperature of the heated lid or is this always a fixed value? For example, if I want to do a reverse transcriptase PCR where I...
02 February 2016 4,646 8 View
Hi. I've been using fin clips from zebrafish for genomic DNA extraction, followed by PCR, and then ExoI and TSAP treatment. I then send these off for sequencing. So far I've been using this method...
02 February 2016 2,094 4 View
I did a PCR for cDNA from 3 different tissues, 2 of which do not express my gene of interest. While only the predicted tissue showed the band for my gene of interest, the negative controls for...
01 January 2016 5,677 10 View
I am planning to do an in situ to determine if a gene of interest is expressed in the intestine. I have done in situs in larval zebrafish but never in adults/adult tissue. Can anyone point me...
01 January 2016 8,610 7 View
I am trying to obtain cDNA from RNA samples from 5 different tissues. Currently, I am doing the following: 1ug RNA + random oligomer (50uM) + water followed by 10mins at 70 degrees C. Then I add...
12 December 2015 4,551 10 View
I want to do a qPCR to detect expression levels for a gene transcript in zebrafish. I have wild types and homozygous mutants for this mutation and it is a nonsense mutation. Since the mRNA...
10 October 2015 2,955 3 View
I am cloning out a fragment, using SacII, containing the ORF of my interest and cloning it into a SacII site in the multiple cloning site of my vector. The insert can obviously orientate itself in...
04 April 2015 9,665 10 View
I am struggling with a three way gateway cloning for several transgenes that I have been trying to clone for a month. I am using three entry clones and one destination vector with the LR Clonase...
08 August 2014 2,255 8 View
I've been working with some very large constructs which I've obtained via the multi-gateway cloning technique. I have successfully obtained these from mini preps but when I turn to midi preps, I...
02 February 2014 6,744 13 View
I have been trying to clone a ~2.5kb insert into an ~8kb vector. I have been given varying advice and to make things worse, the cloning hasn't been working. I want to start all over and identify...
08 August 2013 5,527 6 View
I want to pour out LB agar plates for transforming some constructs that have both ampicillin and chloramphenicol resistance markers. The concentration of ampicillin is 50mg/ml and that of...
08 August 2013 1,677 4 View
Hi. I am trying to access this publication titled "Growth in Fishes" by H Dutta, 1994. Unfortunately, our dept. does not have access. Would it be possible for anyone to help out with this, please?...
01 January 1970 6,229 3 View
