Hi guys. It's my first time doing RNA extractions. I'm trying to extract from soybean seed using the Promega kit. I think I've managed to get some RNA but judging from gel pic the quality is not good. Nanodrop readings (A260/A280; A260/A230) are as follows a: (2.8;1.7), b: (2.6;0.77), c: (3.2;1.7); d: (2.7;2) & e: 2.7; 1.5 (from left to right lane of pic). Any suggestions on how I can improve? Thank you.
Dear Gugulethu Joy Makatini
Pls. find the attached files, may help in your research, good luck.
https://www.takara.co.kr/file/manual/pdf/9769_e.v1309Da.pdf
http://link.springer.com/article/10.1007/BF02668374
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0059270
http://www.biotechniques.com/multimedia/archive/00004/BTN_A_05386BM05_O_4355a.pdf
http://bmcgenomics.biomedcentral.com/articles/10.1186/1471-2164-11-136
http://www.sciencedirect.com/science/article/pii/S2215016116000066
Regards
Prof. Houda Kawas
Hi Gugulethu Joy Makatini,
I think your samples are partially degraded, you may need to do it in a RNase-free environment. 260/280 are really high, maybe the RNA are too concentrated. You could dilute it and test again. For 260/230, it seems like there is contamination of organic solvents. Try to purify it again with ethanol.
Hope it may helps,
Guangqi
Make sure everything that your RNA is going to come into contact with is RNAse free. Tips, gloves, tubes, pipetters...everything. Try getting some RNAse Away or something similar and wipe it down. Prep your RNA in a different location, and with different pipetters (i.e. don't prep RNA samples in a spot you'd use to isolate DNA...if you are able).
Following the useful comments from others I can just add a few things. RNA definitely looks degraded. It is vital that you freeze your sample immediately when you take it- straight into liquid nitrogen. I would recommend an additional clean-up step for these samples- we use a Lithium Chloride precipitation which works well.
- Badges
- Science method
- Similar topics
- Chemistry
- Biochemistry
- Biochemical Methods
- Protein Purification Techniques