Navigation Links
Oragene and DNA sequencing with ABI PRISM

J. Chartier
DNA Genotek Inc., Ottawa, ON, Canada

DNA from Oragene is reliably sequenced using the ABI PRISM 377 DNA Sequencer.


Introduction
Blood leukocytes are the traditional source of DNA for molecular testing, but collection is invasive and specimens require infectious precautions. Buccal swabs are a less-invasive method of collecting DNA for sequencing applications such as HLA typing and prelingual deafness screening (ref. 1, 2) but they tend to have an appreciable failure rate (ref. 3). Oragene is a non-invasive DNA self-collection kit from saliva that provides significantly higher DNA yields than buccal swabs (ref. 4). The purpose of this study was to investigate the suitability of DNA from Oragene for DNA sequencing.


Materials and Methods
DNA collection
Saliva samples were collected from 5 donors. Collection and purification of DNA was carried out according to protocols supplied with the Oragene kit.

Primer design
PCR primers for the human Thymidylate Synthase (TYMS) gene were designed based on the publicly available DNA sequence (GenBank accession no. AP001178). The primers generate a 560-bp fragment. Table 1 shows the primer sequences.

DNA Sequencing
Oragene-purified DNA from each of the 5 samples was used as the template for PCR with the TS 560 primers. The PCR products were sequenced in both directions by Cortec DNA Service Laboratories (Kingston, ON) using an ABI PRISM 377 DNA Sequencer (Applied Biosystems) and the DYEnamic ET Dye Terminator Kit (Amersham Biosciences). Prior to sequencing, the PCR products were purified using the microCLEAN DNA clean-up reagent (Microzone).


Results
DNA sequencing results were aligned using CLUSTAL W (Version 1.83), a multiple sequence alignment program. All 5 samples correctly aligned to the original human Thymidylate Synthase gene sequence. Figure 1 shows a representative ABI PRISM sequencing read-out.


Discussion and Conclusions
Oragene is a non-invasive and reliable method of collecting DNA from saliva. DNA from Oragene works well for DNA sequencing with the ABI PRISM 377 DNA Sequencer.


References
1. Liu, C., Chu, C., Lee, Y., Shi, Y., and M. Lin. (2004). Identification of two novel HLA-DRB1 alleles, DRB1*0903 and DRB1*1145. Tissue Antigens. 64: 99101.

2. Sugata, A. et al. (2002). High-throughput screening for GJB2 mutations its clinical application to genetic testing in prelingual deafness screening for GJB2 mutations. Auris, Nasus, Larynx. 29: 231-239.

3. Elit, L., Jack, E., Kwan, E., Baigal, G., and S. Narod. (2001). A unique BRCA1 mutation identified in Mongolia. International Journal of Gynecological Cancer. 11: 241-243.

4. DNA Genotek. (2004). DNA Yield with Oragene. WP002 Rev 1.5 Sept 04.


back to top
'"/>

Source:


Page: All 1 2

Related biology technology :

1. DNA Stability with Oragene
2. Oragene and Whole Genome Amplification with GenomiPhi
3. DNA Yield with Oragene
4. Oragene is compatible with TaqMan SNP genotyping
5. Serial Analysis of Gene Expression Using ABI PRISM DNA Sequencers and BigDye Terminators
6. Total RNA Purification from Cultured Cells Using the ABI PRISM 6700 Automated Nucleic Acid Workstation and Total RNA Lysis Reagents
7. Rapid, High Throughput Mutation Discovery Using Fluorescent SSCP on the ABI PRISM 3100 and 3100Avant Genetic Analyzers
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:2/12/2016)... 12, 2016 Biocom, the association representing the ... group of San Diego companies to ... 2016 Precision Medicine Advocacy Fly-In. Biocom Fly-In participants had the ... Administration (FDA), the Centers for Medicare and Medicaid Services (CMS), ... Diego U.S. Representatives Susan Davis and Scott ...
(Date:2/12/2016)... 12, 2016 MedGenome,s Commitment ... Scientific Understanding of Complex Diseases Such as Cancer, ... --> --> MedGenome, ... and a leading provider of genomics research services ... to the GenomeAsia 100K consortium as a founding ...
(Date:2/11/2016)... Germany and GERMANTOWN, Maryland ... ; Frankfurt Prime Standard: QIA) today announced the introduction ... for gene expression profiling, expanding QIAGEN,s portfolio of Sample ... enable researchers to select from over 20,000 human genes ... interactions between genes, cellular phenotypes and disease processes. ...
(Date:2/11/2016)... New York, NY (PRWEB) , ... ... ... International (BPI), a business-to-business publication dedicated to delivering cutting-edge information focused on ... GE Healthcare Life Sciences to become a premier sponsor of the 2016 ...
Breaking Biology Technology:
(Date:2/3/2016)... , February 3, 2016 ... market research report "Automated Fingerprint Identification System Market by ... Search), Application (Banking & Finance, Government, Healthcare, and Transportation) ... MarketsandMarkets, the market is expected to be worth USD ... 21.0% between 2015 and 2020. The transformation and technology ...
(Date:2/3/2016)... , Feb. 3, 2016 ... the addition of the "Emotion Detection ... Machine Learning, and Others), Software Tools (Facial ... Areas, End Users,and Regions - Global forecast ... --> http://www.researchandmarkets.com/research/d8zjcd/emotion_detection ) has ...
(Date:2/2/2016)... This BCC Research report provides a ... the recent advances in high throughput ‘omic platforms ... forward. Includes forecast through 2019. Use ... opportunities that exist in the bioinformatic market. Analyze ... well as IT and bioinformatics service providers. Analyze ...
Breaking Biology News(10 mins):