Navigation Links
Comparison of PCR Kleen Spin Columns to Traditional Methods for Purification of PCR Products Prior to Sequencing, Rev A

Jolanta Kolodziejek, MSc, and Professor Norbert Nowotny, PhD, Clinical Virology Group, Institute of Virology, University of Veterinary Medicine, Vienna , Veterinaerplatz 1, A 1210 Vienna, Austria


Introduction
The purity of the DNA template employed in sequencing PCR (cycle sequencing) is the most critical factor in nucleotide sequencing. The presence of contaminants such as enzymes (nucleases, polymerases), salts, primers, primer- dimers , nucleotides (dNTPs or ddNTPs), ethanol, and others decre ases the quality of the sequencing scans significantly. Primer- or RNA-contaminated samples exhibit a high background and lead to misinterpretation of sequencing data. Contamination with nucleases, salts, nucleotides, and ethanol decreases read length and signal intensity and sometimes results in unre a dable sequence data.

We tested Bio-Rads PCR Kleen spin columns for purification of PCR products prior to sequencing PCR in two different experiments, and compared the resulting nucleotide sequences obtained from these samples with the sequences obtained after purification by traditional methods.


Methods
Experiment 1. Comparison of Effectiveness of Purification Method Prior to Sequencing PCR
A 510 bp RT-PCR product (100 l) was prepared using Borna disease virus as template, and three of four equal-volume aliquots of the PCR product were purified by different methods prior to sequencing. The fourth aliquot was left as an unpurified control.

The first aliquot of the PCR product was purified using PCR Kleen spin columns according to the instructions. The DNA f rom the other two aliquots was recovered from agarose sli
'"/>

Source:


Page: All 1 2 3 4

Related biology technology :

1. PCR Performance Comparisons Between pfuturbo and Taq DNA Polymerases
2. Performance Comparisons of Commercial RT-PCR Systems
3. A Comparison Study of Lipid Tranfection Reagents in A549, NIH 3T3 and COS-7 Cell Lines
4. Comparison of Growth Techniques and Media for the Purpose of Plasmid Isolation from E. coli Using the Eppendorf Perfectprep Plasmid Mini Kit
5. TripleMaster PCR System: Direct Comparison with Competitors
6. Quantitative Measurement of Cell Proliferation Using the BrdU ELISA: A Comparison Between Colorimetric and Chemiluminescent Detection
7. Comparison of different protein quantitation methods
8. Comparison of Different Methods of mRNA Quantitation
9. Bio-Plex Cytokine Immunoassays and ELISA: Comparison of Two Methodologies in Testing Samples From Asthmatic and Healthy Children, Rev A
10. Performance Comparison of the Experion Automated Electrophoresis System and a Competing Automated System for Protein Analysis, Rev A
11. Separation and Comparison of Proteins From Virulent and Nonvirulent Strains of the Fish Pathogen Flavobacterium psychrophilum, Using a 2-D Electrophoretic Approach
Post Your Comments:
(Date:8/28/2014)... 28, 2014  Next month, executives from clinical trial marketing ... events beginning with Patient-Centered Clinical Trials 2014 , to ... Boston , September 4-5. Patient recruitment experts Bonnie ... Fleishman will share insights on the benefits of employing ... tactics – from media to mobile apps – can be ...
(Date:8/28/2014)... Sterlitech is proud to announce that ... stable of products . These additional products ... include membrane filters with surface charges. , ... for our membrane process testing equipment, and correspondingly, a ... membranes,” explains Sterlitech President Mark Spatz. “The addition ...
(Date:8/28/2014)... Exciting new work by a Florida State University research ... can take your temperature, emit white light, and convert ... molecule looks like a butterfly. , Biwu Ma, associate ... in the FAMU-FSU College of Engineering, created the molecule ... continued to discover that his creation has many other ...
(Date:8/28/2014)... Vt. (PRWEB) August 28, 2014 ... testing around the world is available from Strategic ... diagnostics companies. “ Industrial Microbiology Market Review, Fourth ... Industrial Market ” (IMMR—4) tracks and compares microbiology ... in North America, Europe and Asia, and forecasts ...
Breaking Biology Technology:BBK Worldwide Leads Sessions at Key September Events 2Sterlitech Corporation Enhances Its Selection of Flat Sheet Membrane Filters 2Novel 'butterfly' molecule could build new sensors, photoenergy conversion devices 22 Billion Industrial Microbiology Tests Conducted for Product Quality and Safety Worldwide 22 Billion Industrial Microbiology Tests Conducted for Product Quality and Safety Worldwide 3
... 29 Cadence Pharmaceuticals,Inc. (Nasdaq: CADX ... and commercializing proprietary product candidates principally,for use ... the company,s,President and CEO Theodore Schroeder will ... 6 at 1:30pm Eastern Time (10:30am Pacific ...
... U.S. Attorney,s, Office for Eastern District of Pennsylvania and ... Settles State Cases with Connecticut and ... (Nasdaq:,CEPH) announced today it has finalized a previously announced ... Eastern District of,Pennsylvania, the U.S. Department of Justice and ...
... Calif., Sept. 29 Poniard,Pharmaceuticals, Inc. (Nasdaq: PARD ... Jerry McMahon, Ph.D., chairman and CEO,will present at the ... 9 a.m. Eastern Time at Le Parker Meridien in ... overview, including a discussion,of the development of picoplatin, the ...
Cached Biology Technology:Cadence Pharmaceuticals' CEO Theodore Schroeder to Present at the JMP Securities Third Annual Healthcare Focus Conference 2Cephalon Concludes All Outstanding Federal and State Government Investigations into the Company's Sales and Promotional Practices 2Cephalon Concludes All Outstanding Federal and State Government Investigations into the Company's Sales and Promotional Practices 3Cephalon Concludes All Outstanding Federal and State Government Investigations into the Company's Sales and Promotional Practices 4Poniard Pharmaceuticals to Present at the JMP Securities Healthcare Focus Conference 2
(Date:9/1/2014)... 1, 2014, 10:20 AM EDT) It may be ... foods over unhealthy higher-calorie foods, according to new research ... Research Center on Aging (USDA HNRCA) at Tufts University ... the journal Nutrition & Diabetes , a brain ... it is possible to reverse the addictive power of ...
(Date:9/1/2014)... ago, electron microscopy revealed the presence of tiny blob-like ... But scientists still don,t know what they do ... believed to be crucial to the life of a ... disease treatment., In the Journal of Cell Biology ... from various backgrounds, from biophysics to cell biology, to ...
(Date:9/1/2014)... Section of the XVIII International Botanical Congress took place ... held every six years and it is where the ... fungi and plants get together to debate and update ... This is the primary product of the meeting, the ... which was published in 2012. The other important product ...
Breaking Biology News(10 mins):Training your brain to prefer healthy foods 2Scientists call for investigation of mysterious cloud-like collections in cells 2Week-long meeting on naming algae, fungi, and plants recorded for posterity 2
... Works to Strengthen Security for Anyone with ... Sept. 17 Today a,Southwest Airlines pilot ... be screened by,the Transportation Security Administration (TSA) ... demonstration project called,SecureScreen will utilize biometrics to ...
... $24 million in federal funding to establish the University ... CEIN), which will help researchers design safer and more ... to be housed at the California NanoSystems Institute (CNSI) ... nanomaterials on life forms and the interactions of these ...
... Foundation (NSF) and the Environmental Protection Agency (EPA) have ... Implications of NanoTechnology (CEINT) to explore the potential ecological ... as a million times smaller than the head of ... objects made from the same material. These unusual properties ...
Cached Biology News:Southwest Pilots Launch TSA Demonstration Project to Test Biometric Screening 2UCLA, partners establish new center on environmental effects of nanotechnology 2UCLA, partners establish new center on environmental effects of nanotechnology 3UCLA, partners establish new center on environmental effects of nanotechnology 4Duke to lead new NSF, EPA center to study the environmental implications of nanotechnology 2Duke to lead new NSF, EPA center to study the environmental implications of nanotechnology 3
... IT siRNA Tracker Intracellular Localization Kit provides ... and deliver siRNA, of your design, in ... vitro tracking experiments. Subcellular localization and functional ... monitored following introduction of the labeled siRNA ...
... Beacon 2000 instrument can be used as ... 2000 Data Manager Software provides the means ... from the instrument. These data can then ... spreadsheets, such as Microsoft Excel or Lotus ...
... for the study of protein:protein interactions from ... that interact with GST•Tag™, S•Tag™, or ... using affinity resins for the tags, and ... filter. The complexes are eluted with sample ...
... Microarray Kit (V2), 22K is useful for investigating ... Each microarray contains content derived from the ATH1 ... Research (TIGR) and represents 21,500 genes. ... x 3? glass slides each containing an identical ...
Biology Products: