08 August 2018 2 2K Report

I am using Agilent miRNA first strand synthesis kit (cat#600036) for preparing mi cDNA from total RNA. A universal primer is provided with the kit sequence of which is not given. i want to study expression of some miRNAs using SYBR green baseq qPCR. It is suggested to use the sequence of particular miRNA as the forward primer. What other considerations should be kept in mind while designing such primers as Tm for miRNA sequence in usually low ?

More Bhoomika Sharma's questions See All
No signal in SYBR Green QPCR for miRNA quantification?

Hello everyone ! I am using mirVana miRNA isolation kit for the isolation of total RNA containing small RNA. Further i used Agilent first strand synthesis kit(cat#600036) for the synthesis of...

07 August 2018 800 5 View

Problem with PCR amplification?

i tried gradient PCR for a new set of primers and got the amplification. However, when i tried running the same set of primers on a single annealing temperature, it showed no or weak...

10 November 2016 9,289 14 View

Can anyone tell how to troubleshoot no amplification in PCR?

I am trying to amplify my genes of interest.Though I got the amplified product of the desired size on my first attempt , i am not able to get the amplification again. What might be the problem and...

07 August 2016 9,484 3 View

Problem with RNA Isolation using mirVAna kit?

I have been using mirVana miRNA isolation kit for total rna isolation and had a good experience using that. Recently, i am facing problem with the kit. 280/230 ratio and concentrations are too low...

03 April 2016 5,063 3 View

Problem with RNA isolation from Breast tissue ?

I am trying to isolate RNA from Breast tumor and adjacent normal tissues (stored in RNAlater at -20 C)  using Trizol method. After adding isopropanol , I am getting a transparent droplet like...

02 March 2016 1,479 3 View

Why do I get peaks in NTC in qPCR ?

I am performing real time pcr for ACTB. I am getting a ct value of 17-20 in my samples. However , i am getting a peak in NTC too with a ct value around 33-35. Also the dissociation curve for NTC...

10 November 2015 3,796 8 View

Can anyone help explain double bands in PCR?

I am getting two bands below 100 bp in addition to the gapdh amplicon . What might be the reason for these bands? Lane 2- positive control lane 3-6 - samples  lane 8- negative control

06 July 2015 4,145 6 View

What factors should be considered in miRNA -target prediction?

I am trying to predict miRNAs against the gene of interest using online tools like Targetscan, miRanda ,miRsystem and DIANA micro T- CDS. As these tools produce a long list of miRNAs that might...

06 July 2015 8,395 6 View

How can I select a good endogenous control for miRNA qPCR?

I have to perform qPCR for miRNA isolated from colorectal tumor and adjacent normal tissue samples. Is U6 snRNA a good endogenous control for qPCR?

05 June 2015 3,690 9 View

Do you know how I can process TCGA data?

I have downloaded the miRNA seq data for colon adenocarcinoma from TCGA. If I would like to see the expression of a particular miRNA across all the samples, how should we proceed?

03 April 2015 5,573 1 View

Similar questions and discussions
Anyone having idea about VN primer for miRNA primer design ?

How to design VN primer to attach with universal reverse primer

05 August 2024 2,116 3 View

Could dyes amplify the spectrum of light to a specific wavelength?

I am interested to know the behavior of dyes toward light. Specifically, Blue dyes re-emit the spectrum, especially from the green zone (known as principal in LED lamps, and blue dyes are known...

05 August 2024 3,290 1 View

What is the problem with these tissue culture plants?

All plants are green but some of these plants becomes yellow. I did not found any reason. Please help me to find out the real problem.

01 August 2024 589 4 View

Transfection in HEK293T cells?

Dear All, I am trying to transfect a pCDNA3.1 vector containing my gene of interest. The purpose is to figure out the localization of the protein of interest. I have fused the protein with GFP on...

31 July 2024 9,892 4 View

How to retain the a GFP tagged gene expression in stable cell line?

Hello , I established a stable cell line expressing GFP tagged to a centrosomal gene having G418 drug selection marker. I validated the stable line by IFA and Western blotting, results are fine....

29 July 2024 5,007 0 View

AGILENT HPLC 1100 SERIES , any expert available please?

AGILENT HPLC 1100 SERIES operation assistance needed.

25 July 2024 10,045 2 View

Can I please ask why my samples from anaerobic bioreactor giving me different size PCR product even after multiple runs?

Hi everyone, I have extracted DNA from a biogas bioreactor using Qiagen kit and prep cDNA library then used this library as template to optimize primers for qPCR (taken from papers). Some of the...

23 July 2024 1,329 5 View

How to quantify the fluorescence intensity of only the merge area of an image using ImageJ software?

Dear All: I have to quantify the fluorescence intensity of cells that are fluorescent to two different markers, so i'm interested in finding out if this is possible. Previously I have...

22 July 2024 8,770 1 View

What are the key risks associated with foodborne pathogens in imported leafy greens, the impact on public health and effect on regulatory framework?

I am currently writing a systematic review

15 July 2024 9,176 1 View

How do i go about my research which is about the integration of solar and wind power to produce green hydrogen?

Let me say I have a solar plant in operation and I want to build up a wind power plant, and later make a hybrid so that I could power the electrolyser as load for green hydrogen production.

14 July 2024 9,547 4 View

Our partners will collect data and use cookies for ad personalization and measurement. Learn how we and our ad partner Google, collect and use data. Agree & close