When sequencing PCR products, how much volume of amplicon is required?

More Rais Ahmed's questions See All

If in mann whitnet test U value comes in thousands..U=1435, then is it correct??

if U= 1435 in mann whitney test, then is it correct?? i mean U value may be in thousands???

09 October 2016 7,779 0 View

If mann whitney test value falls in thousands then what it means??

i got mann whitney test value U=1402, i am confused about this so large value. this value is right? i mean if value comes in thousands then no need to worry or there is any problem??? please

09 October 2016 2,891 2 View

Is there any association of body weight with paratuberculosis in cattle?

hello anyone who has studied any article that explains the relationship of body weight with onset of paratuberculosis (Johne disease) in cattle..please suggest

07 August 2016 9,339 0 View

How to write qualitative data in Graph pad prism software?

hello, i want to enter qualitative data like yes or No OR positive or Negative in Graph pad prism software but how to enter??? please suggest if anyone knows..

07 August 2016 6,397 1 View

Is there any research article published that indicate bacterial DNA degradation/survival in soil?

dear all scientists, i am in search of an article that contain information about Bacterial DNA degradation/ survival in soil with effect of different temp, pH, humidity etc...please inform if...

07 August 2016 2,867 2 View

DNA sequencing tel you about specie?

hello, if DNA sequencing is done on genus specific primes, then the sequencing of amplicon can tel us about the specific specie???

31 December 2015 8,112 0 View

I am facing problem in optimizing real time PCR...?

i am facing problem in real time pcr optimization, i am using Bio-Rad system CFX96. i got different peaks on screen. but could not got sigmoid curve. when i run these samples on gel i got a clear...

11 December 2015 9,068 6 View

Any advice on the problem in optimization of real time PCR?

i am facing problem in real time PCR optimization. i am using Bio Rad system CFX96. I found Tm of primers 54C in gradient PCR. then i run real time PCR but no peaks are seen but i found clear...

10 November 2015 722 4 View

What are controls in PCR? if we want to get positive control, then how can we get? i mean comercially available? or from where we got?

molecular biology

07 August 2015 4,307 5 View

Which companies make bacterial DNA ? i mean for PCR if we need positive control then which companies supply it?

molecular biology, microbiology, genetics, biotechnology

07 August 2015 6,645 2 View

Can I base on reverse DNA sequences to perform alignment, convert to amino acids and GenBank submission?

I have reverse sequences (AB1 format), can I base on reverse DNA sequences to perform nucleotide alignment, convert nucleotides to amino acids and deposit the sequence in GenBank database?

11 August 2024 5,138 1 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

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

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

Does anyone have issues using Prepman Ultra reagent for MicroSeq ID bacterial, fungal and yeast sample preparation?

I have been attempting to extract DNA from Bacterial, Fungal and Yeast banked samples (>1e7 cells) using Prepman Ultra reagent and I seem to be struggling to obtain a sequence. Although the...

01 August 2024 2,079 0 View

I'm optimizing a tetra-ARMS PCR protocol with amplicon sizes: 165bp, 123bp and 93bp. Why, in the gel, only 165bp is visible while I'm expecting all 3?

It's an end-point PCR protocol. I'm using 1.5% agarose gel with SyBR Safe dye and TBE as a running buffer, visualization on BioRad XR+ system. I was primarily thinking of primer efficiency,...

01 August 2024 4,673 4 View

PCR showing no bands - Master mix and primers didn't mix?

Hello everyone, I performed a PCR yesterday, and the results showed no bands on the gel. Of course, I probably missed some crucial steps, like adding my samples to the PCR strips themselves, for...

31 July 2024 2,406 6 View

Dimensions of an MJ Research 96-well alpha module?

Can anyone with an MJ research / BioRad PCR machine from ~2010 or earlier tell me the external measurements (LxWxH) of the removable standard PCR alpha modules that can be removed from a PTC200 or...

30 July 2024 2,867 0 View

How to retain amplicon yield after Ampure bead cleanup, following a 2-stage PCR protocol?

Hello all, I have been trying to follow a 2-stage PCR protocol used to amplify barcodes of a large yeast library, as per Nyugen et al. (2022) -...

30 July 2024 841 2 View

Should the amount of DNA input used for ChIP-seq library preparation be matched between the control and experimental groups?

Hi all. As a beginner in ChIP-seq experiments, I hope you understand that the following questions might be somewhat basic. I am planning to perform ChIP-seq or MeDIP-seq analysis to investigate...

28 July 2024 6,938 1 View

Rais Ahmed

i think about 20 micro litre is sufficient???

Irfan Khattak

Dear Rais

Different sequencing companies have different volume requirements so it would be better to ask your sequencing company about the exact volume of PCR product required

Arfat Yousaf Shaheen

Dr. Rais!

Our ABI company require 20uL of PCR product which is sufficient for sequencing but it should be > 20uL.

thanks for answering

Syed A Ali

Hello Rais,

The rule is 10 ng of PCR product for every 100 bp of DNA fragment.