HOW can i minimize the number of hits made by the designed primer to only the intended sequence?

i have designed a primer but i found it gives 200 hits over than the intended sequence (5 of them in the organism that i work with). this give poor pcr results. i want to avoid this but i don,t...

08 September 2018 6,894 3 View

Could any one help me in designing accurate primers for the following sequence?

i had designed primers for two sequences using blast tools in NCBI but they gave poor amplification lower than the ladder after gradient pcr i want any one to check them and help me in designing...

08 September 2018 7,829 4 View

How to calculate delta CT for calibrator in qpcr in case-control study ?

i do a study for a gene expression in HCC patients compared to healthy person using real time pcr. i have analyzed ct for gene of interest and house keeping gene in each sample either HCC or...

06 July 2018 6,703 13 View

Is it useful for doing conventional PCR before REAL TIME PCR to be sure of the product size and primer annealing?

i will do a real time PCR for miRNA after revrse transcription. iam afraid from bad results and wasting mu kits which is expensive. i want to know whether it is possible to do conventional pcr and...

02 March 2018 3,804 12 View

What is the best PCR condition (PCR mix and cycles and temperature ) for amplification of a WNT10A?

I have used the following procedures but am still having strong primer dimer bands while the desired product is absent. The cycling conditions for PCR program that i used were 5 min at 95C for...

02 March 2018 5,925 6 View

How can i make pedigree file for using in haploview software ?

i want to make haplotype and linkage analysis. i found that the most common programme is the haploview. but i don,t know how to make pedigree or linkage or premakeped file to use in the haploview....

02 March 2016 917 0 View

What is the difference betwwen gold and green open access?

i want to choose the green open access because it is free but i want to know every thing about it

01 February 2016 3,075 4 View

Is it better ti use triazole or total RNA or miRNA isolation kit to isolate miRNA (less than 200 nt) from serum or plasma?

10 November 2015 7,260 8 View

Is qRT-PCR one step or two step better for miRNA?

10 November 2015 3,207 3 View

How to design primers for microRNA?

I WANT TO DESIGN primers for microRNA

10 November 2015 2,101 1 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

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

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

Does RNAiMAX transfection reagent work in serum free medium?

I diluted siRNA and RNAiMAX in opti-MEM and added to the cells which they were in the growth medium. Is it a right way? or should I culture cells in the opti-MEM medium for a while and not in...

26 February 2021 10,041 3 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