Navigation Links
Easy Cosmid DNA Purification with StrataPrep Kits


Miniprep and midiprep quantities of pure high-kb DNA

Jeff Braman Scott Basehore
Stratagene

The StrataPrep plasmid miniprep and midiprep kits are designed to isolate up to 20 and 350 g, respectively, of high-purity plasmid DNA from a single spin cup.1,2 These kits can also easily purify very large plasmids, including cosmid DNA.

To demonstrate cosmid purification, a SuperCos I clone containing a 40-kb insert from human chromosome 11 was isolated from two, 100-ml overnight cultures using the StrataPrep EF plasmid midiprep kit. The cosmid DNA yield from the two cultures was 45 and 51 g, respectively. The A260/A280 nm ratios were 1.8 and 1.9, and the A260/A240 nm ratios were both 1.7, indicating an absence of protein, SDS, and EDTA contaminants.3 Contaminating chromosomal DNA was also absent. The resulting cosmid DNA was digested to completion with EcoR I (Figure 1).

Fig.1

Cosmid DNA can be also be isolated from 1.5-ml overnight cultures using the StrataPrep miniprep kit. The cosmid DNA is completely digested with restriction enzymes such as EcoR I (Figure 2).1

Fig.2

The midiprep purified cosmid DNA was subjected to automated fluorescent sequencing, a very sensitive application demanding pure template DNA. A representation of the high-quality sequence is shown in Figure 3. The read length was approximately 630 bases with only four sequence ambiguities.

Fig.3

The StrataPrep plasmid miniprep and midiprep kits both produce high yields of all sizes of DNA from 2.9-kb plasmids to 45-kb cosmids. Both have rapid protocols using a modified alkaline lysis method to yield quality sequence with automated or manual methods. The amount of purified DNA required determines which kit to use. StrataPrep purified cosmid DNA can serve as the source material for subclone library construction used in shotgun sequencing projects.4

REFERENCES
  1. Basehore, S. and Braman, J. (1998) Strategies 12: 5-7.
  2. Braman, J. and Basehore, S. (1997) Strategies 10: 84-86.
  3. Held, P. (1998) Pharmaceutical Laboratory, February issue.
  4. Wilson, R. K. and Mardis, E. (1997) In B. Birren, et al (Eds.), Genome Analysis, A Laboratory Manual. pp. 397-403, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.

'"/>

Source:


Page: All 1 2

Related biology technology :

1. Isolation of Cosmids using Eppendorfs Perfectprep Plasmid Mini Kit
2. High-Level Protein Expression, One-Column Purification, and FLAG Epitope Tagging in E. coli
3. Simple Purification of DNA from Plasmid Minipreps, PCR Amplifications and Agarose Gels
4. Yeast Protein Production System Features High Yields and One-Step Purification
5. Expression and Purification of Recombinant Proteins That Have Native Amino Acid Sequence
6. MinElute PCR Purification Kit
7. QIAquick PCR Purification Kit
8. Microarray Target Purification Kit and the Microarray Target Synthesis Kit (T7)
9. Double-Affinity Purification of GST-His Tagged Proteins Using Vivapure 8-to-96 Well Cobalt-Chelate Kit
10. Ultra-Fast Purification of Small Nucleic Acids
11. RNAClean - RNA & cDNA in vitro Reaction Purification
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:5/19/2016)... ... ... Anton Paar USA, located in Ashland, Virginia is pleased to announce that construction ... a third office building to the current facilities. , Growth is nothing new. ... with office space adjacent to the previous main building. Through remodels and new construction, ...
(Date:5/19/2016)... 2016 - I dati saranno ... il 52 ° Congresso della ... - Le conclusioni dello studio indicano un tasso ... di cui il 90% presenta una d urata della ... Il settantadue per cento dei pazienti ha riscontrato un beneficio ...
(Date:5/18/2016)... ... May 18, 2016 , ... Every day, more than ... asthma complications.* Costing more than $56 billion in direct costs annually, asthma remains ... too many, the suffering associated with uncontrolled asthma can be overwhelmingly disproportionate and ...
(Date:5/17/2016)... Saudi Arabia (PRWEB) , ... May 17, 2016 ... ... the biopharmaceutical industry, and BioSmartSA, a healthcare consultancy based in Saudi Arabia, have ... of diagnostic services to healthcare providers in the Kingdom of Saudi Arabia (KSA). ...
Breaking Biology Technology:
(Date:4/14/2016)... , April 14, 2016 ... Malware Detection, today announced the appointment of Eyal ... new role. Goldwerger,s leadership appointment comes at ... heels of the deployment of its platform at several ... biometric technology, which discerns unique cognitive and physiological factors, ...
(Date:3/31/2016)... 2016  Genomics firm Nabsys has completed a financial ... Bready , M.D., who returned to the company in ... leadership team, including Chief Technology Officer, John Oliver ... Nurnberg and Vice President of Software and Informatics, ... Dr. Bready served as CEO of Nabsys from ...
(Date:3/22/2016)... , March 22, 2016 ... research report "Electronic Sensors Market for Consumer Industry by ... & Others), Application (Communication & IT, Entertainment, ... - Global Forecast to 2022", published by ... is expected to reach USD 26.76 Billion ...
Breaking Biology News(10 mins):