Oct 17, 2021
Evagreen Dye Fluorescence Verification
- 1Chung Shan Medical University
- CSMU_Taiwan
Protocol Citation: Chia-Hsien Shih 2021. Evagreen Dye Fluorescence Verification. protocols.io https://dx.doi.org/10.17504/protocols.io.by5ppy5n
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it’s working
Created: October 17, 2021
Last Modified: October 17, 2021
Protocol Integer ID: 54159
Abstract
This protocol is to establish the relationship between Evagreen Dye fluorescent intensity and DNA concentration.
Preparation
Preparation
Take 30 µL of 10µM ssDNA (T4 ligation primer) into a new eppendorf
Dilute 10µM ssDNA into 1µM :
Take 3 µL of 10µM ssDNA and 27 µL RNase-free water into a new eppendorf
Dilute 1µM ssDNA into 100nM :
Take 3 µL of 1µM ssDNA and 27 µL RNase-free water into a new eppendorf
Dilute 100nM ssDNA into 10nM :
Take 3 µL of 100nM ssDNA and 27 µL RNase-free water into a new eppendorf
Dilute 10nM ssDNA into 1nM :
Take 3 µL of 10nM ssDNA and 27 µL RNase-free water into a new eppendorf
Dilute 1nM ssDNA into 100pM :
Take 3 µL of 1nM ssDNA and 27 µL RNase-free water into a new eppendorf
Dilute 100pM ssDNA into 10pM :
Take 3 µL of 100pM ssDNA and 27 µL RNase-free water into a new eppendorf
Add 1.5 µL of 20X evagree dye into each eppendorf.
Measure
Measure
Respectively load 20 µL of 10µM,1µM, 100nM, 10nM, 1nM, 100pM and 10pM ssDNA into each well
Measure the fluorescence excitation and emission intensity