How to make working concentration of around 2000 uM of Temozolomide such that it contains DMSO of 0.22 % or less for use in cell culture?

I am working on a drug called Temozolomide in my cell culture experiments. I am having trouble in preparing working concentrations of this drug from 200 to 2000 uM as the drug is soluble in DMSO...

06 July 2019 7,188 3 View

Can an antibody that can be used for Immuno-cytochemistry can also be used for Immuno-fluorescence ?

I am planning to order antibodies for my experiment which can be used for ChIP, Western blot, Immuno-fluorescence (IF) and Immuno-histochemistry. Some suppliers mention their product to be...

01 February 2018 1,609 5 View

Can higher dosage of antibiotics added to the mammalian cell cultures affect the cells?

We normally add a very low concentration of antibiotics like Penicillin, Streptomycin, Chloramphenicol or Tetracycline or a mixture of them in the culture plate to avoid bacterial growth in the...

31 December 2017 6,473 1 View

Why Salmon or Herring sperm DNA is used for blocking the membrane and target DNA in Southern blotting?

We all know that Herring sperm DNA or Salmon sperm DNA is used for blocking non-specific binding in Southern blotting. My question is what makes them the best choice for blocking non-specific...

31 December 2017 8,960 1 View

What may happen if our antibiotic vial gets contaminated with a little amount of cell culture?

While treating the adherent mammalian cells in a T-25 flask with an antibiotic (just after changing the media) can we use the same tip for adding the same antibiotic to another flask? Is it ok?...

31 December 2017 5,719 7 View

Why my lower MW bands are not visible in SDS PAGE?

03 April 2017 6,797 20 View

What is the maximum size of amplicon can an NEB Taq Polymerase (normal one) amplify?

What I have heard is the normal Taq Polymerase which we commonly use can amplify a fragment upto 1.5 kb. But yesterday someone told me that the normal Taq Pol can amplify a fragment upto 3kb...

08 September 2016 8,900 3 View

What is the Gibbs free energy of a normal PCR reaction?

I want to actually find out whether a PCR reaction is spontaneous or non-spontaneous and if it is spontaneous how far it is spontaneous?

08 September 2016 1,742 2 View

Is it possible that the plant showing positive for a transgene in T2, the offsprings of the plant in T3 show negetive for the transgene?

I did touchdown PCR using gene specific primer targeting lectin gene inserted into the cotton genome by isolating DNA from T2 plants.The PCR showed positive result in one of the event plant. The...

08 September 2016 5,415 5 View

If one of my PCR component is some how contaminated then is it possible that I get bands in some of my blanks and that to faint and not in all blanks?

Actually I was checking whether my Primer stock is contaminated or not. So I kept 6 blanks and one positive. Out of 6 blanks I got faint bands in 2 of the blanks(all components except template)....

03 April 2016 1,496 4 View

How do I increase the yield of my PCR product?

Hello, We would like to increase the yield of our PCR product. We are running a series of PCR reactions that is targeting ~1.1kb sequence. We begin each reaction with ~400pg of template DNA...

02 March 2021 4,029 3 View

PACYC PCR not working after several attempts. Would anyone have any suggestions?

So, I have been trying to run a pACYC PCR which will be used later on for a Gibson Assembly. However the PCR is not working. I have already tried gradient PCR and changing extension time; however...

02 March 2021 1,146 2 View

Does anyone know HRM channel in a 2 plex QIAGEN Rotor- Gene Q qPCR can be used as a separate channel?

I am going to have 3 different probes in my qPCR work that I am going to do. But I realized that the machine we have in the lab is a Rotor-Gene Q 2plex HRM Platform, saying it has green, yellow,...

01 March 2021 8,544 1 View

How to address this issue in the analysis of Real-Time PCR results?

To dear Researchers, I was analyzing a series of concentration for estimation of Real-Time PCR efficiency. The concentration was 1:10. I used MS-excel to evaluate Slope. The result of slope was -8...

01 March 2021 8,683 4 View

No title

Does anyone have the experience of using Taq Man probes in the QIAGEN Rotar- Gene qPCR machine?

01 March 2021 5,311 1 View

MiRNA qpcr problem?

Dear All, mirna primer showing some problem in the melting curve? any idea why? As attached is the melting curve. The forward sequence is obtained from miRBase and reverse primer is universal.

28 February 2021 5,008 4 View

Runs of nucleotides in Sanger sequencing?

I performed site directed mutagenesis, transformation, and then I sent out plasmids for Sanger sequencing and found out that there is extension of DNA just before the stop codon. I am not sure...

27 February 2021 547 3 View

Can thermal cycler machines be problematic during PCR?

Hi, my question is about the heating of thermal cycler machines and I hope some of you had experienced a similar thing previously. There are two thermal cycler machines in the lab(BioRad) and for...

26 February 2021 4,777 4 View

Calculating volume of DNA required from DNA concentration?

Hi I am a bit confused. They are asking me to find out the volume of DNA required in ul (a total of 30-100 ng for genomic DNA) from the DNA concentration in the nanodrop reading which was 404.8...

26 February 2021 5,029 2 View

Has anyone used the Illumina ITS1 primer sets?

Hi everyone, Illumina provides a list of primers to amplify with high taxonomic coverage the ITS1 region for further fungal sequencing, but I cannot find the exact amount necessary of each...

25 February 2021 6,969 3 View