What impact would be there on RNA-SEQ experiments if we do not use any RNA Spike-In controls (like ERCC or SIRVs)?

Hi everyone, if I do not use spike-in RNA isoform controls for a customized RNA sequencing experiment (not the whole transcriptome, just based on few specific genes we are interesting), so is...

01 February 2020 8,059 2 View

Can I add "enzyme-based RNAse Inhibitors" to the extracted FFPE RNA elutions to avoid any possible RNA degradation by RNAses?

Hi I am using RNAse Inhibitors (applied biosystems), which are enzyme-based and are meant to add during cDNA synthesis phase as per its manufacturer policy (1ul to each 20 ul RT reaction). But,...

04 May 2019 9,014 2 View

Is there any undesirable effect of bacterial RNA load of normal intestinal flora present on human colon samples for the gene expression analyses?

It is a known fact now that interactions between bacterial (normal flora) and colon cancer cells influence the transcription profile of host cells or enterocytes. So, how can I avoid or minimize...

09 October 2018 6,099 1 View

Can we pool together the same type of samples in qPCR for delta-delta ct method?

Is it worthy to pool all the samples together belong to a same group, but originated from different animals of a same species, to save time and resources (reagents)? For example; From my negative...

05 June 2018 8,555 7 View

Does "No Primer Dimer Formation" guarantee the absolute utilization of entire primer content (F & R both) in a PCR reaction?

Dear all, in a PCR (conventional & realtime both), if we achieve our desired product (GOI) and along with, there is also no primer dimer (PD) formation seen on the gel and melt curve, does...

10 November 2017 4,434 4 View

How to get rid of PCR amplification artifacts while using the Taq DNA polymerase in PCR to prepare the libraries for NGS applications?

Taq DP is very well known to induce Indels and single base substitutions etc. So if any polymorphism or genetic variation detected later on, through any NGS platform, what makes us sure that these...

04 May 2017 4,099 4 View

How an Asymmetric qPCR (unequal forward and reverse primers) results in an improved Melt Curve?

A low concentration of a primer than the other one in high concentration or even in volume often exhibits good melt curve appearances. What could be the reasons behind this? Is this dependent on...

10 November 2016 4,951 3 View

The frequency of distribution of CRISPR elements among Gram -ve and Gram +ve bacteria almost the same or more in G +ve than G -ve or vice versa?

10 November 2016 4,244 0 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

Gel electrophoresis result for total RNA samples, 1.5 Agarose gel was used, How to improve the integrity of RNA?

Gel electrophoresis, RNA degradation, RNA extraction from fresh tissue

02 March 2021 5,433 5 View

Does anyone have a recommendation for yeast reporter gene plasmid/protocol?

Hi, could anyone recommend a plasmid and/or protocol for reporter gene assay in S. cerevisiae? I want to assess the effect of growth conditions on a transcription factor, so I want to clone it`s...

01 March 2021 210 1 View

Ester cleavage conditions?

Hi all, I need to cleave the ester in Ethyl 2-amino-1H-imidazole-5-carboxylate to make the carboxylic acid and retrieve the product from solution to use in downstream applications of linking to...

01 March 2021 8,766 3 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

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

01 March 2021 5,311 1 View

How vaccines of covid 19 work?

Millions ask how vaccines of covid 19 be successful and safe, its here to explain generally how vaccine of pfizer work ,mRNA vaccines teach our cells how to make a protein that will trigger an...

28 February 2021 2,723 4 View

Expression cloning by using cDNA,do I need to modify the cDNA first?

I am going to have a expression cloning of mammalian gene by using shuttle plasmid to transforming the E.coli However I don't know I should only inserting the Coding sequence ,or I can...

28 February 2021 5,440 3 View

Problem with shRNA transfection. Why are only two plates getting contaminated instead of all?

I transfected my LNCaP-WT cells with 3 shRNA plus their NTC two weeks ago and split two puromycin selected cell plates on Friday last week(Feb 26). I checked for GFP in the cells, and they all...

28 February 2021 4,949 3 View