SEQanswers

Go Back   SEQanswers > Introductions



Similar Threads
Thread Thread Starter Forum Replies Last Post
PubMed: Methylome analysis using MeDIP-seq with low DNA concentrations. Newsbot! Literature Watch 1 04-25-2017 04:06 AM

Reply
 
Thread Tools
Old 10-15-2017, 06:22 AM   #1
John D
Junior Member
 
Location: Australia

Join Date: Oct 2017
Posts: 1
Smile DNA concentrations

Hi

hoping someone can assist - lawyer working on DNA case

If the extracted DNA is .25 nanograms per microlitre how would the biologist achieve .5 nanograms per microliter for purposes of then amplifying with the amplification kit???

Is it as simple as adding 2 microlitres to the solution???? I don't get how that increased the amount of DNA to be amplified as the concentration seems to remain the same??

Thanks

John
John D is offline   Reply With Quote
Old 10-15-2017, 10:47 AM   #2
Ola
Member
 
Location: Sweden

Join Date: Aug 2011
Posts: 30
Default

Hi,

adding a larger volume would make it easier to amplify even though the concentration is the same since you then will have more template molecules. It is also possible to amplify from less than 0.5 nanogram (one copy of the human genome weighs ~0.006 ng).
Ola is offline   Reply With Quote
Old 10-15-2017, 12:12 PM   #3
nucacidhunter
Senior Member
 
Location: Melbourne

Join Date: Jan 2013
Posts: 1,144
Default

To double the contration, volume of DNA solution need to be halfed. Three options:

1- dry down the solution with speedyvac or similar device. This will increase the salts concentration which might be detrimental for some downstream applications.

2- using clean up columns or beads in which DNA binds to a solid support and then eluting bound DNA with less volume of buffer. Some DNA will be lost in the process.

3- using Amicon Ultra Centrifugal filters which allows liquid and small molecules to pass through a filter and DNA stays behind.
nucacidhunter is offline   Reply With Quote
Old 10-16-2017, 06:14 AM   #4
seqtechno1
Member
 
Location: U.S.

Join Date: May 2017
Posts: 12
Default concentrating DNA

Do a concentration and purification using AMPure XP magnetic beads (they are SPRI beads, you can google how they work if interested) and concentrate and purify the DNA using these and a magnetic stand/rack. The trick is you will also need some sort of re suspension buffer which typically contains PEG I think? Its hard to tell b/c most of them are proprietary and come in disposable kits used for sequencing.
seqtechno1 is offline   Reply With Quote
Reply

Thread Tools

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is Off




All times are GMT -8. The time now is 08:52 PM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2018, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO