Mace, E.S. and Buhariwalla, K.K. and Buhariwalla, H.K. and Crouch, J.H. (2003) A high-throughput DNA extraction protocol for tropical molecular breeding programs. Plant Molecular Biology Reporter, 21 (4). p. 459.
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Abstract
Liquid handling robotics and capillary electrophoresis genetic analyzers now offer high-throughput solutions for 2 of the 4 key steps in PCR-based DNA marker-assisted fingerprinting (DNA extraction, PCR amplification, electrophoresis, data analysis). Thus, DNA extraction remains the most significant bottleneck at the bench for large-scale applications in plant breeding and germplasm characterization. We report on a rapid and low-cost method for relatively high-throughput extraction of high-quality DNA from young and mature leaves of sorghum, pearl millet, chickpea, groundnut, and pigeonpea. The procedure uses a modified CTAB/β-mercaptoethanol method for DNA extraction in a 96-well plate. The quantity and quality of the DNA extracted per sample is adequate for more than 1000 PCR reactions. A relatively high throughput of 96–384 samples per person per day can be achieved, depending on the crop. A major timesaving aspect of the protocol is the absence of a manual sample-grinding step. Finally, the cost is a magnitude lower than commercial plate-based kits, and, as such, is likely to have substantial application in tropical molecular breeding programs.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | DNA isolation; High throughput; Molecular breeding; PCR |
| Author Affiliation: | Hermitage Research Station, 604 Yangan Road, 4370, Warwick, QLD, Australia |
| Subjects: | Statistics and Experimentation Crop Improvement |
| Divisions: | Other Crops |
| Depositing User: | Ms Ishrath Durafsha |
| Date Deposited: | 23 Nov 2013 14:16 |
| Last Modified: | 23 Nov 2013 14:16 |
| Official URL: | http://dx.doi.org/10.1007/BF02772596 |
| URI: | http://eprints.icrisat.ac.in/id/eprint/12158 |
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