If liquid nitrogen is not avaliable, what can we use in DNA extraction?
Please indicate the source of DNA extraction to determine whether you really need liquid nitrogen or not. In most cases, one can incubate a sample at -80 degree C for 20 min to precipitate DNA.
Liquid Nitrogen is a cryogenic fluid that helps to break the cell wall and prevent phenolic oxidation
Alternative protocol
A Simple Method for DNA Extraction from Mature Date Palm Leaves: Impact of Sand Grinding and Composition of Lysis Buffer
Ibrahim A. Arif, Mohammad A. Bakir, Haseeb A. Khan,* Anis Ahamed, Ahmad H. Al Farhan, Ali A. Al Homaidan, Mohammad Al Sadoon, Ali H. Bahkali, and Mohammad Shobrak
DNA Extraction
Fresh leaf of date palm (100 mg) was placed in a sterile mortar. Sterile sand (50 mg) and 500 μL of lysis buffer (Table 1; lysis buffers A to E) were added separately to the sterile mortar. Leaf sample was finely crushed using mortar and pestle and allowed to dry at room temperature for about 5 min. Crushed leaf sample with sand (100 mg) was transferred into a 1.5 mL eppendorf tube. The same lysis buffer (1,000 μL) that was used for grinding the leaf was added to the tube and vortexed briefly. The tube was then kept in a water bath at 60 °C for 30 min. After mixing by brief vortex, the tube was centrifuged at 9,500 g for 5 min. An aliquot of supernatant (200 μL) was transferred to a new tube, taking care to avoid carryover of any dirt or debris. An equal volume (200 μL) of chloroform: isoamyl alcohol (24:1) was added and the tube was shaken gently top to bottom for 5 min followed by centrifugation at 9,500 g for 5 min. The supernatant (200 μL) was transferred to a new tube and sodium acetate (3.0 M; 20 μL) plus cold isopropanol (500 μL) were added gently and the tube was kept in the freezer for 5 min followed by centrifugation at 11,500 g for 10 min. The resulting supernatant was discarded and 500 μL of 70% cold ethanol was added and vortexed briefly. After centrifugation at 7,000 g for 5 min, the supernatant was discarded and the tube contents were air dried at room temperature. DNA was eluted with 100 μL of TE buffer and kept at 4 °C for further use. Experiments were conducted with 4 individual replicates.
Depending on what you are extracting DNA from, you may be able to use a different protocol that doesn't require freezing the sample at all. In my lab, we extract DNA from mouse tail snips by overnight incubation with proteinase K, then add salt and chloroform, followed by freezing and thawing (and then more steps). We used to freeze on dry ice, but now just put the samples in the -80 for about 20 minutes. Perhaps dry ice or a -80 freezer would work for you. If you don't have those, maybe you could just put the samples in a -20 long enough for them to freeze solid. I don't know if this will work, so I would recommend trying it on some unimportant samples first. Good luck.
Hi: You can use a DNA extraction kit that not required liquid nitrogen, Or you can use freeze dryer the sample and grind with ceramic or iron balls in the hight speed shaker.
Thank you all.
Actually i am extracting DNA from dried sample of Prosopis Juliflora by CTAB method. but when i check DNA spectrophotomer reading it is near to 1.08. I have to use RAPD markers but in literature the reading on spectrophotometer should be 1.8.
So, i am checking other possibilities to improve DNA extraction, DNA extraction kit is good but we are not allowed to use it (manual work).
plz comment
Actually, you don't need liquid N in DNA extraction. I think it is not the reason for getting poor yield of DNA. You must improve your extraction protocol, your sample, the solutions you used in your protocol.
I agree with Nibras, it is definitely something to do with the way you extract the DNA and not liquid Nitrogen. I personally do RNA extractions from leaf samples and I love using dry ice, easy to handle and works best. your method you need to look at every step: 1. starting from the sample condition and amount (how much sample are you using?) 2. Then efficient grinding, to make sure maximum cell contents are released. 3. Optimum incubation and centrifugation times at different steps as you go from removing RNA, proteins and salts from your sample during the extraction process. encouraging comment: CTAB is very efficient in terms of yield and quality, but only downside is that it is a bit laborious/time consuming..
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