Navigation Links
Quantitation of DNA for Automated Sequencing Using the VersaFluor Fluorometer

standard curve was established for quantification of DNA concentrations ranging from 1005,000 g/ml by plotting seven points of varying DNA concentrations made by serial dilution of control calf thymus DNA. All dilutions were made in 2 ml of 1 g/ml Hoechst 33258 dye and included 0 ng DNA as a blank and 10,000 ng DNA as the upper range. The values were recorded in relative fluorescence units (RFUs) and were plotted on a graph with the x axis = DNA concentration (ng) and the y axis = RFU (Figure 2). The slope of the line was determined to be 1.0.

By employing the equation y = mx+b, we established the value of b to be equal to 0. Therefore, by setting the upper range value at 10,000 RFU using 10 l of 1 mg/ml calf thymus DNA and using 2 l of the sample DNA in 2 ml of the 1 g/ml Hoechst dye, the RFU reading is equal to ng/ml for our sample. We are then able to calculate the volume of DNA needed to obtain a 250 femtamolar concentration per reaction.


Results and Discussion
Using the Excel II sequencing kit (Epicenter) and quantifying our DNA with the VersaFluor fluorometer, we have been able to achieve highly readable and reproducible sequencing runs of up to 1,200 bp with the LiCor 4200 automated sequencer. This has enabled us to sequence the entire 2.2 Kb cDNA for Xenopus hypoxia inducible factor, xHIF, in several runs, and will allow us to sequence our genomic clone easily and efficiently.

The correct concentration of DNA in each reaction is the key to obtaining good reads on the LiCor 4200. This entails precise quantitation of the DNA. The VersaFluor fluorometer has proven to be easy to calibrate and provides excellent and reproducible results.

This work is
'"/>

Source:


Page: All 1 2 3

Related biology technology :

1. Quantitation of Cabergoline at Extremely Low Plasma Concentrations with a Triple Quadrupole Mass Spectrometer
2. Bioanalytical Method Intraday Validation for the Quantitation of Paroxetine in Bovine Plasma using the TSQ Quantum Mass Spectrometer
3. Quantitation of the Protein MIA as a Marker for Chondrocytes in Research Samples
4. Comparison of Different Methods of mRNA Quantitation
5. Quantitation of RNA for Hybridization Blots Using the VersaFluor Fluorometer
6. Quantitation of Lymphangiogenesis Using the iCycler iQ Real-Time PCR Detection System and Scorpions Detection System, Rev A
7. Quantitation of GAD67 Gene Expression in Prefrontal Cortex of Schizophrenia Patients Using the iCycler iQ Detection System and Molecular Beacons, Rev A
8. Benchmark Plus Microplate Reader: Quantitation of Protein Concentration Using Two Different Colorimetric Assay Kits, Rev A
9. A Robust LC/MS/MS Method for the Quantitation of Very Long Chain Fatty Acids for Research into the Characterization of Peroxisomal Disorders
10. A new LC/MS/MS Research Method for Rapid Quantitation of Five Immunosuppressant Drugs, including Mycophenolic Acid (MPA)
11. Separation and Quantitation of R and S Enantiomers using Normal Phase Chromatography on an API 3000 LC/MS/MS System
Post Your Comments:
(Date:5/23/2013)... MARINO, Calif. , May 23, 2013 /PRNewswire/ ... today a strategic alliance with DAK Renewable Research ... the corn oil yield. VG Energy is a ... (OTC Pink: VGLS). VG Energy,s exclusively ... to increase the oil yield in plants, possessing ...
(Date:5/23/2013)... EXTON, Pa. , May 23, 2013 /PRNewswire/ ... leading research and advisory firms for specialized biopharmaceutical ... gastroenterologists, hepatologists, and infectious disease specialists reported that ... warehousing patients (e.g., intentionally delaying treatment) in anticipation ... physicians than six months ago, when only 6 ...
(Date:5/23/2013)... Md. , May 23, 2013 Northwest ... biotechnology company developing DCVax ® personalized immune therapies ... use its exhibit at this year,s American Society Of ... to highlight its leadership role in immune therapy.  ... The ASCO annual meeting is the pre-eminent conference focused ...
(Date:5/23/2013)... SAN DIEGO , May 23, 2013  Verenium ... biotechnology company focused on the development and commercialization of ... , Chief Financial Officer, will present at two ... Mr. Black will present at the Second Annual ... at the Grand Hyatt Hotel in New York City.  ...
Breaking Biology Technology:VG Energy Enters into a Strategic Alliance with DAK Renewable Research for LipidMax Field Trials 2VG Energy Enters into a Strategic Alliance with DAK Renewable Research for LipidMax Field Trials 3The Majority of Physicians that Treat Hepatitis C Virus (HCV) Have Begun "Warehousing" and Preparing Their HCV Patients for the Next Generation of HCV Treatments 2The Majority of Physicians that Treat Hepatitis C Virus (HCV) Have Begun "Warehousing" and Preparing Their HCV Patients for the Next Generation of HCV Treatments 3NW Bio Exhibit at the Upcoming ASCO Meeting to Highlight its Leadership Role in Immune Therapy for Cancer 2NW Bio Exhibit at the Upcoming ASCO Meeting to Highlight its Leadership Role in Immune Therapy for Cancer 3Verenium To Present At Two Upcoming Conferences 2Verenium To Present At Two Upcoming Conferences 3
... of Missouri officials announced today a five-year agreement with ... research and training in nanomedicine and nanotechnology. "We ... affiliation of 16 institutions of higher education in Tamil ... research. "K.S.R. has the same commitment to deliver quality ...
... Mich., Oct. 11, 2011 New England Biolabs, Inc. ... signed a distribution agreement whereby NEB will distribute a ... the amplification of DNA from single cells or extremely ... Genome Amplification (WGA) Kit is the first product available ...
... 11, 2011 Roche ( SIX: RO, ... announced today an agreement to license several technologies developed ... Arizona State University and Dr. Colin Nuckolls of the ... revolutionary DNA sequencing system. The licensed technologies include specialized ...
Cached Biology Technology:MU signs academic agreement with K.S.R. Educational Institutions of India 2New England Biolabs and Rubicon Genomics Enter into Distribution Agreement for Single Cell Amplification Products 2Roche Licenses Nanopore Sequencing Technologies from Arizona State University and Columbia University for Rapid, Affordable DNA Sequencing 2Roche Licenses Nanopore Sequencing Technologies from Arizona State University and Columbia University for Rapid, Affordable DNA Sequencing 3Roche Licenses Nanopore Sequencing Technologies from Arizona State University and Columbia University for Rapid, Affordable DNA Sequencing 4
(Date:5/23/2013)... Wis. In an age when microbial pathogens are ... tamp down infection, a team of Wisconsin scientists has ... curbing the bacteria that cause staph infections. , Writing ... Society , a group led by University of Wisconsin-Madison ... with the "quorum sensing" behavior of Staphylococcus aureus ...
(Date:5/22/2013)... to improve durability in fuel cell powered buses, ... discovered links between electrode degradation processes and bus ... effects of electrode degradation stressors in the operating ... , The findings of the study, led by ... in a long-term study at Burnaby-based Ballard Power ...
(Date:5/22/2013)... 2013)The omega 3 fatty acids in fish oil have ... so that the American Heart Association currently recommends eating ... fatty varieties rich in omega 3s. However, the mechanism ... a new study, scientists led by Jason R. Carter ... by providing evidence that fish oil might specifically counteract ...
Breaking Biology News(10 mins):University of Wisconsin chemists find new compounds to curb staph infection 2Tests lead to doubling of fuel cell life 2Fish oil may help the heart beat mental stress 2
... discovered molecular mechanism helps control the amount and effectiveness of ... that is produced by the body,s own nerve cells. ... Nature Neuroscience . The lead author on the study is ... the University of Washington (UW). The senior author is Dr. ...
... outdoor "laboratory" on Ohio State University,s campus shows that naturally ... ecological services as will wetlands planted by humans. Researchers ... experimental marshes on the campus, one that was planted in ... colonize plant and animal life on its own. The 2 ...
... ITHACA, N.Y. Apathetic aphids which become ... and alarms sent by fellow aphids become easy ... to researchers at the Boyce Thompson Institute for Plant ... Academy of Sciences , Aug. 3, 2010.) The ...
Cached Biology News:Newly discovered mechanism controls levels and efficacy of a marijuana-like substance in the brain 2Newly discovered mechanism controls levels and efficacy of a marijuana-like substance in the brain 3Planted, unplanted man-made wetlands are similar at year 15, and function as effective carbon sinks 2Planted, unplanted man-made wetlands are similar at year 15, and function as effective carbon sinks 3Planted, unplanted man-made wetlands are similar at year 15, and function as effective carbon sinks 4Apathetic aphids with attitude become easier prey for ladybugs 2
... The Helios gene gun low-pressure system, 220-240 ... biomaterials into cells. This handheld device employs an ... other biomaterial-coated gold microcarriers from the inner wall ... cells. This system can be used for in ...
... Hybridization Wash Pack provides optimized blocking ... Preparation Quality Clarifier: Ready-to-use wash solutions ... SigmaScreen(tm) APS Coated Slides for Microarrays ... Hybridization Blocking Buffer: 1L • Microarray ...
Mouse polyclonal antibody raised against a partial recombinant MHC2TA. NCBI Entrez Gene ID = 4261...
Mouse monoclonal antibody raised against a partial recombinant JRKL. NCBI Entrez Gene ID = JRKL...
Biology Products: