|Register||Search||Today's Posts||Mark Forums Read|
|Agarose Gel Electrophoresis Forum Agarose Gel Electrophoresis Forum. A forum to discuss electrophoresis staining, troubleshooting, and development of films for DNA and RNA gels.|
| ||LinkBack||Thread Tools||Display Modes|
DNA gel electrophoresis: protein contamination?
Hi im kinda new here so if this is in the wrong section, then thats ok.
Well, what i an trying to find using 1.2% Agarose gel electrophoresis is to prove that there is protein contamination in my plant extracts (my plant extracts collect a month ago were freezed dried after collection before being grounded using a motar & pestle and finally was stored around 25 degrees (room temperature)).
This is so when i know there is protein contamination (which is what i want before finding some biophenols using High performance liquid chromatography.)
So what i did was to use 1ul microliter Dnase, rnase and proteinase K for each of my plant extracts. Positive control was the plant extract and was diluted (in 1/20) using distilled water. Negative control was Nuclease free water. Both contained 6x loading dye. A gene ladder was used in my sample to only know if the gel was working.
Note: None of the sample underwent purification process before loading the gel with the samples. Also note that the same samples tested on a biophotometer had a 230/260nm ratio of 1.01 for DNA/RNA purity and the 230 absorbance was low (about .8A) for the 1/20 sample
To get straight to the point, this is what each well contained
Well 1 : Empty
Well 2 : 3ul gene ladder with 2ul dye and 2ul Nuclease free water (NFW).
Well 3 : 4ul NFW and 2ul loading dye
Well 4: 7ul Diluted 1/20 sample and 2ul loading dye and 6ul of NFW
Wells 5-7 : 7 ul of diluted 1/20 sample with either 1ul of DNase, rnase and Pro K in each well, each well had 2ul dye and 6ul of NFW
well 8 had 1ul of DNase,Rnase and pro K (total 3ul) in the well with 6ul of diluted 1/20 sample, 2ul loading dye and 4ul of NFW
When the gel is made, only one band appear (which was the gene ladder), on the wells, and they were bright. However there is a faint brightness around the well. Could that mean something or that i have to make another agarose gel with the samples again?
Also if anyone can link my to a better protocol, that would be gladly accepted.
|contamination , dna , electrophoresis , gel , protein|
|Thread||Thread Starter||Forum||Replies||Last Post|
|Human Cytome Project - Update 24 Jan. 2005||Peter Van Osta||Cell Biology and Cell Culture||1||08-01-2010 02:18 PM|
|Human Cytome Project - Update 6 Jan. 2005||Peter Van Osta||Cell Biology and Cell Culture||0||01-06-2005 10:18 AM|
|New Saccharomyces Sequences 11/27/04||Mike Cherry||Yeast Forum||0||11-28-2004 11:39 PM|
|New Saccharomyces Sequences 05/19/04||SGD Sequences||Yeast Forum||0||05-23-2004 04:06 PM|
|New Saccharomyces Sequences 09/10/03||SGD Sequences||Yeast Forum||0||09-11-2003 01:33 AM|