11 ug of total plasmid isn't too bad for a miniprep in my opinion. How did the nanodrop spectra look like. Did you have 260nm/280nm values of around 1.8? Anything too much off would suggest contamination, repeat prep with a new kit box. I never had luck sequencing those guys. You should be able to estimate the concentration by agarose gel. You will find the concentration of  the DNA fragments in the standard in the manual. Use the standard to get an estimation of your plasmid concentration.

Also, did you double check the sequencing primer?

If you don't see anything on an agarose gel you most likely don't have any plasmid from the miniprep. How much DNA are you adding to the gel? I would recommend adding 200-500 ng of the plasmid to the gel in order to see a very bright band. 

Dear Justin,

If it is a low copy number plasmid, you will get a fairly consistent concentration after miniprep. If you would like to concentrate it further you can apply the elution of one column to the next till you get a high enough concentration. I have done this myself and it works. Secondly, if the sequencing facility is mentioning absence of DNA, maybe you could check if your tubes (in which you store or send the DNA) are nuclease free. Since, even if the DNA is degraded, it will give a reading on the nanodrop. It is not a problem with the machine, but rather a caviat we can cross check by running your samples on gels.

P.S. you could also try a midi or maxi prep.

Regards,

Dear Justin

• I agree with Cornelia: 10ug of plasmid DNA from a mini prep falls within the realms of what you would expect
• In terms of Nano drop 'anticipated recovery' remember that measurements @ 260nm are not specific to nucleic acids: They actually measure (pi) electron excitation in aromatic rings and will thus 'aromatic contaminants'. Trizol for manual extractions and guanidinium salts in the lysis buffer of kit based mini prep methods
• Thus Nano drop has a habit of overestimating DNA recovery and thus I routinely check by agarose gel to verify ACTUAL recovery
• As mentioned by Justin 200ng to 500ng measured by Nanodrop is an optimal amount to load onto an agarose gel to obtain high signal & low noise without saturation (which happens at > 1ug)
• I will mention in passing that I tend to recommend to students 100ng of total DNA estimated from a gel for each kb of total construct for Big Dye sequencing
• Finally, if you require concentration of your purified plasmid DNA, precipitation with 1/10 vol 3M Sodium acetate pH4-5 & 3 x volumes of molecular grade ethanol, preferably with 1ul of 1mg/ml to 20mg/ml glycogen is effective
• Details of this method are elaborated in a prior research gate answer of mine:

https://www.researchgate.net/post/how_to_make_concentration_high_of_DNA_extracting_from_PCR_Product#view=578c9799ed99e132b905cde3      

Hi Justin,

This a link to a nanodrop bulletin explaining 260/280 and 260/230 ratios which may give you further information how to deal with your problem:

http://www.nanodrop.com/Library/T009-NanoDrop%201000-&-NanoDrop%208000-Nucleic-Acid-Purity-Ratios.pdf

Please give us an update once you solved your problem!

Before sending your plasmid DNA for sequencing, you should have examined your agarose gel. Well let me tell you the quantity 11micro gram means nothing as it could be a highly sheared DNA also and the sequencer won't find anything to sequence. Check your sample and repeat the experiment.