Seeing 10 ng DNA in a single band is no problem for agarose gels, stained after the run with 0.5 microgram per ml ethidium bromide, and photographed on a normal UV-transilluminator. Lower amounts are sometimes  more complicated, and very narrow and sharp bands need to be produced during electrophoresis.

By the way: Against all rumours: EtBr is much less toxic than most people suppose, and mutagenicity is really low There are no mutants seen in the Ames test without treating the substance with microsome fractions, and very few mutants even after treatment.

Generally speaking, it's difficult to visualize less than 20 ng of DNA per lane. This can be affected, however, by lane width (smaller lane, more concentration), the sensitivity of your imaging system and the concentration of DNA stain you add.

Hello milind!

can you give us some details about the downtream use of your DNA?

if your DNA is a pcr product, normally you could see it in gel using small lanes (as Jordan suggested) but if you are dealing with genomic or fragmented DNA (where you expect a smear) the quantity has to be much higher to be visualized.

sometimes is better to perform optimisation steps first and, once your protocol is reproducible, you can use your DNA for your downstream experiments without checking it on gel if the quantity is the limitating factor.

With most staining molecules (EtBr, Eurosafe, GelRed) the limit of detection is approximately 5 ng/lane if you use a well large 3-4 mm. If  you want to keep most of your DNA for downstream applications, try to load 10 ng (1 ul).

Hello Filippo,

It is a genomic DNA and I would like to check, if it is there or not. and its concentration is very low. so I would like know that what is the minimum concentration of DNA is required to visualize on agarose gel.

Thanx  Enrico sir...

Dear Milind,

how did you quantify your DNA to say it is 10 ng/ul? If you use a spectrophotometer, it is likely that the concentration is much lower since extraction of DNA brings with it also nucleotides and ribonucleotides, that absorbe at 260 like DNA (slightly muche more to say the truth). So if after misuration you think to have 10 ng/ul, is is likely that the concentration of genomic DNA is lower, let's say1-2 ng/ul. If you do not have issues to use large volumes, load 10-15 ul of your genomic DNA.

ok sir .. I will try it...thanx..

Great Answer Enrico. It is very likely the case here.

I agree with Enrico. if you have more quantity of your sample, then you can use more volume to load in gel.

At least 5 nG of NA is required to light up as a band, is the standard that I used in my PhD work. Is that close? NANOTECHNOLOGY is on the way to improving that. We have an introductory course in Nanotechnology being taught for the first time this Fall 2014 at Erie Community College-North Campus. 

Hi Milind,

Nowadays, some labs have replaced EtBr with non-mutagenic and non-cytotoxic DNA dye. GelRed is one of the examples. The amount of DNA needed for one to visualize the DNA on the gel may also vary depending on the dye we use. Attached is a picture showing gel staining with EtBr or GelRed (lanes: 200 ng, 100 ng, 50 ng and 25 ng) [picture was from Cambridge Bioscience website].

The minimum amount of DNA detectable by ethidium bromide on a 3-mm-thick gel and a 5-mm-wide lane is 1 ng.

Seeing 10 ng DNA in a single band is no problem for agarose gels, stained after the run with 0.5 microgram per ml ethidium bromide, and photographed on a normal UV-transilluminator. Lower amounts are sometimes  more complicated, and very narrow and sharp bands need to be produced during electrophoresis.

By the way: Against all rumours: EtBr is much less toxic than most people suppose, and mutagenicity is really low There are no mutants seen in the Ames test without treating the substance with microsome fractions, and very few mutants even after treatment.

Generally we can see 8-10 ng DNA on gel as a band.

we see up to 10ng with gel red

With EtBr, I would say my answer perfectly coincides with Dr. Wöstemeyer from Jena University. As an advice, I suggest to use fresh EtBr solutions. In my experience, old preparations tend to render unsatisfactory staining and then, underestimation of the DNA amount.   

I would see one can see as little as 10 ng on an agarose gel

I aggree with previous answers : fresh BET gives better visualisation, and also bath of BET solution can give better result than melting BET directliy in the gel.

(for example we use Smart Ladder at lab, as you can see the smallest amount visible in the ladder is 20 ng (200bp band in 5 mm lane width), with others ladders, you can see 10 ng)  http://www.eurogentec.com/uploads/TDS-MW-1700-10.pdf -

https://norgenbiotek.com/display-product.php?ID=45

If your dna length is short, you may have problems to visualize it on a melted ETBr agarose gel (EtBr is charged +, so it migrates inversely than DNA). So as said before, use a fresh bath. Idealy, i would use polyacrylamide gel and a more sensitive product for revelation.

The DNA visualized on the gel varies and depends on the dye. The minimum detectable amount of DNA using ethidium bromide is 1 ng.

10ul of you sample (with 3-5ng/ul ) will be more than enough to be visualized on the gel.

About 25 ng of DNA will give excellent results on agarose gel. Provided if there are no other contaminants.

The amount is actually sufficient to carry out the process provided proper pore size as well as good concentration of agarose gel should be taken into consideration.

10 ng is fine for PCR. It is better to take use 2ul for AGE if your DNA conc is 10..

Hello Milind,

10 ng is the minimum amount of DNA to visualize it on agarose gel. However, to play safe you can start with 20 ng.

regards

Jitendra

I agree, 10-20 ng will be enough... 

begin with 20 ng... 

Hi,

I would also like to add to above very good discussions that if you have less quantity of DNA and you want to use it further for your experiments then I suggest the you first increase the concentration and quantity of your DNA by doing PCR using your purified DNA as a template. 

Regards,

How much is visible depends also on transiluminator and fluorescent dye you are going to use. UV is for EtBr, SYBR dyes, however other dyes (there is lots of them) may require blue light - check specifications.

20ng

I agree, 10-20 ng will be enough... 

Staining dye Detection limits

BlueView™ 0.2 µg DNA

Ethidium bromide 1 ng DNA

Methylene blue 5 ng DNA

SYBR® Green I 60 pg DNA

SYBR® Green II 0.5 ng RNA

DAPI 0.5 µg DNA

Acridine Orange 50 ng DNA

50-60 nano gram per microliter is enough for genomic DNA

First of all, if you do ethanol precipitation of your 10 ng DNA to increase its concentration then you can run agarose gel electrophoresis by using 1 micro liter of DNA. It can gives result.

You can use gel red as a dye for the gel ans saves you from ethidium bromide. Easy to use just add the dye when making the gel

ive managed to visualise smaller amounts (5-10ng)

most of the time 50-60 nano gram is enough.

I think 20ng is enough just to visualize

I saw a video from addgene, it recommends to use 10-100 ng DNA/well.

I couldn't get a sharp band with 10 ng DNA (Green viewer, uv) whereas running 100 ng of DNA visualized well.

@Milind Mutnale: 10ng-20ng DNA load per well will do just fine.

Even the DNA concentraction (approximate size is 500bp) is around 0.5 ng/µl, load 5µl on the pore and stained with new made 5000 times diluted SYBR Gold, the band could be seen.

Recently I got clear genomic DNA band using 20ng/micro liter. Picture attached.

Many thanks

I have a similar question.

the concentrations of my DNA samples range between 60ng/ul to 100ng/ul. do I have to dilute these samples in order to obtain clear bands on an agarose gel electrophoresis?

you have both options.

you can use same concentrations.

or

you can dilute it to 25 ng/ul.

prefer second.

It depends on the image linearity of your reader. Initially, do multiple sample readings of both undiluted, and then diluted to 25 ng/ul. Next calculate the total intensities from each method, with SEMs, and compare. If not significantly different, no need to dilute. This must be tested over full concentration range at each point in your gel.

Hi,

First, thanks for the informative answers.

I'm currently facing a similar issue trying to visualise DNA from a 4C library.

I have (Nanodrop) concentrations around 20 ng/µl (diluted in TE buffer). I'm loading a total of 30-40 µl (including 6X loading dye) per lane and I'm not getting any band at all. I have GelRed as dye in a 0.6% gel and have run it in two ways: 100 V for 1.5 h or 15-20 V overnight.

Hi, I have a question. How much loading buffer we use for pcr product and how much for digestion product?