How to denature pre-mRNA for RT-PCR ?

30 October 2013 1 9K Report

mRNA are easy to denature but denaturation of pre-mRNA requires additional steps.

I have tried heating at 70C and 80C for 10 minutes prior to cDNA synthesis, but it hasn't worked yet, can someone suggest another method?

Leszek Blaszczyk

Did you use RNAse H to degrade RNA from RNA:DNA hybride? It helped me.

Badges
Science topic

More Tarun Tyagi's questions See All

Do you think can be any Uranium bearing rocks in Eastern part of Iran and western part of Afghanistan?

I want to know more about Uranium ore deposits in world.

11 August 2024 6,720 0 View

Do you think can be any diamond bearing rocks in Eastern part of Iran and western part of Afghanistan?

I want to know more about diamond ore deposits in world.

11 August 2024 2,167 1 View

What is the difference between mathematical R^4 space and physical 4D unit space?

We assume that the difference is huge and that it is not possible to compare the two spaces. The R^4 mathematical space considers time as an external controller and the space itself is immobile in...

10 August 2024 6,678 14 View

If Banks do not provide credit facility, what are the options available for FPOs and impact on producer’s income?

10 August 2024 8,198 5 View

Controlling for pupil light reflex when analyzing pupil size time course?

I used eye tracking to examine how participants from two different populations (A and B) react to an image. Participants in population A exhibit larger pupil sizes over time, but they also have...

10 August 2024 3,229 0 View

What are a “Farmers Producer Organization” (FPO) and its essential features?

10 August 2024 477 5 View

Strugglling with m6A dot blot any suugesstion ?

I have been doing the m6A dot blot for a while with no improvement, I am extracting the RNA, and I can see the dots although the three biological replicas give a different reading on the memberan...

10 August 2024 8,539 5 View

Do interactions between biosphere, carbon cycle, & water cycle impact global warming & interaction between atmosphere & hydrosphere?

How do interactions between the biosphere, the carbon cycle, and the water cycle impact global warming and interaction between the atmosphere and the hydrosphere?

09 August 2024 3,291 2 View

How to get moment output in Abaqus Standart?

I have input a moment load in module load Abaqus, i put my moment load on the node surface (using reference point). I have define moment in history output and make a set for moment too. But the...

08 August 2024 4,831 4 View

How is energy cycled through the Earth's climate system and how do matter cycle and energy flow through the rock cycle?

08 August 2024 8,162 0 View

Is there a problem with my RNA pellet?

Hello, I am currently having problems with RNA extraction. I am using mouse liver (C57BL6J), and I have extracted RNA from mouse liver before. Before this experiment, my final RNA pellets were...

11 August 2024 7,082 3 View

Strugglling with m6A dot blot any suugesstion ?

I have been doing the m6A dot blot for a while with no improvement, I am extracting the RNA, and I can see the dots although the three biological replicas give a different reading on the memberan...

10 August 2024 8,539 5 View

How to confirm the site-directed mutagenesis result without performing NGS?

I'm cloning a fragment of 3200 nts into plasmid. The cloning was successful, however, 02 amino acids were mutated. Now I want to fix these 02 aa by site-directed mutagenesis technique using...

08 August 2024 4,645 2 View

RNA Extraction Using Hot Borate Method No Longer Working?

I've been performing RNA extraction on cotton petiole tissue for a few months now using the method described in the following paper, a derivative of the typical hot borate method...

08 August 2024 9,882 2 View

I can't see the ssDNA band after performing asymmetric PCR. Is there any way to do this?

After performing symmetric PCR, PCR purification was performed. Afterwards, asymmetric PCR was performed using the PCR purification product as a template, but no ssDNA band was confirmed in the...

08 August 2024 1,668 3 View

Does Anyone have expertise in in vitro transcription and RNA pull down assay?

I am currently working on LncRNA; to know the lncRNA-protein interactions I want to do RNA pull down assay, so I need to design primers with T7 promoter. I need assistance in this regard.

07 August 2024 6,622 1 View

Separation of organic acids-HPLC?

Hello What should be done to separate and identify organic acids in HPC when their RetTime is the same?Like oxalic acid with Propanoic Acid.or acids that have a very close RetTime.

07 August 2024 8,782 3 View

E.coli contamination in human RNA seq data ?

Recently, we observed that 99% of the sequences in our RNA-seq data corresponded to the E. coli genome. Despite multiple DNAse treatments after RNA extraction and ribosomal depletion, we were...

06 August 2024 807 3 View

RNA later for the preservation of RNA in fecal samples at room temperature for one day (37°C)?

I am planning to collect human fecal samples for metatranscriptomic analysis using MGI. These samples are from indigenous people living in a region with high temperatures. I will have access to a...

06 August 2024 1,367 3 View

How much total RNA concentration to be extracted from sorted plasma cells from bone marrow of C57BL/6 mice for RT-PCR ?

i have sorted anti-NP specific plasma cells from bone marrow of C57BL/6 mice at certain times after immunization with variable counts and isolated total RNA using TRIZOL method for RT-PCR using...

05 August 2024 8,835 1 View