Luckau Protocols:Agarose Gel: Difference between revisions
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| width="600" style="background-color: #BCED91;" align="center"| <span style="font-size:32px;"> Agarose Gel Protocol </span> | |||
[[ | |align="left" style="background-color: #9DB68C;" | | ||
<span style="font-size:16px;"> [[User:Tara_K._Luckau | Tara K. Luckau's Home Page]] </span> | |||
[[User:Tara_K._Luckau/Notebook/Team_ConGen | Conservation Genetics Lab Notebook]] | |||
[[Luckau_Protocols | Tara's Protocols]] | |||
* [[Luckau_Protocols:NanoDrop | NanoDrop]] | |||
* [[Luckau_Protocols:Agarose_Gel | Agarose Gel]] | |||
* [[Luckau_Protocols:Low_TE | Low TE]] | |||
* [[Luckau_Protocols:Tris-Cl | Tris-Cl]] | |||
* [[Luckau_Protocols:KCl | KCl]] | |||
|} | |||
==Purpose== | |||
Agarose gels are used to verify the presence and estimate the size of DNA (both genomic and amplified). When loaded into a gel and subjected to an electric current, DNA fragments migrate to the positive terminal. Small DNA fragments migrate more rapidly than large ones. By running a commercially available ladder of fragments of known size, we can estimate the DNA fragment size in our sample of interest. | Agarose gels are used to verify the presence and estimate the size of DNA (both genomic and amplified). When loaded into a gel and subjected to an electric current, DNA fragments migrate to the positive terminal. Small DNA fragments migrate more rapidly than large ones. By running a commercially available ladder of fragments of known size, we can estimate the DNA fragment size in our sample of interest. | ||
To visualize the DNA after electrophoresis, a dye called Gel Red is added during the agarose-making process. The dye binds to the DNA and flouresces when exposed to UV light. | To visualize the DNA after electrophoresis, a dye called Gel Red is added during the agarose-making process. The dye binds to the DNA and flouresces when exposed to UV light. | ||
===Materials To Be Familiar With=== | |||
* Gel Rig, caster tray, combs | * Gel Rig, caster tray, combs | ||
* [[Image:OwlA2Large.jpg]] [[Image:OwlA2Large_draw.jpg|300 px]] | * [[Image:OwlA2Large.jpg]] [[Image:OwlA2Large_draw.jpg|300 px]] | ||
==Protocol== | |||
# Make gel using 1x TAE and agarose | # Make gel using 1x TAE and agarose | ||
#* Use 1% for genomic DNA, 2% for amplified DNA | #* Use 1% for genomic DNA, 2% for amplified DNA | ||
Revision as of 13:59, 30 November 2010
| Agarose Gel Protocol |
Purpose
Agarose gels are used to verify the presence and estimate the size of DNA (both genomic and amplified). When loaded into a gel and subjected to an electric current, DNA fragments migrate to the positive terminal. Small DNA fragments migrate more rapidly than large ones. By running a commercially available ladder of fragments of known size, we can estimate the DNA fragment size in our sample of interest.
To visualize the DNA after electrophoresis, a dye called Gel Red is added during the agarose-making process. The dye binds to the DNA and flouresces when exposed to UV light.
Materials To Be Familiar With
- Gel Rig, caster tray, combs
Protocol
- Make gel using 1x TAE and agarose
- Use 1% for genomic DNA, 2% for amplified DNA
| Gel Rig Size | 1x TAE (mL) | 1.5% agarose (g) | 2% agarose (g) | Gel Red (µL) | Sample volume (µL) | max voltage (V) |
|---|---|---|---|---|---|---|
| small | 50 | _ | 1 | 5 | 4-6 | 80 |
| medium | 130 | _ | 2.6 | 13 | 24-tooth comb: 6-10 | 120 |
| 36-tooth comb: 3-5 | ||||||
| large | 270 | _ | 5.4 | 27 | 24-tooth comb: 6-10 | 170 |
| 36-tooth comb: 3-5 |
