Two-dimensional DNA sampling: a new strategy for assessment of hybridity using molecular markers
2000
Nas, T.M.S. | Toledo, R.S. | Li, Z. | Virmani, S.S. (International Rice Research Inst., Los Banos, Laguna (Philippines). Plant Breeding, Genetics and Biochemistry Div.)
A strategy, two-dimensional DNA sampling, was developed for seed purity assessment using amplified fragment length polymorphism (AFLP). Artificially impure populations of the hybrids Magat (IR62828 A /IR29723-143-3-2-1) and Mestizo (IR58025A/IR34686-179-1-2-1) were generated by planting ten row x ten column grow-outs in seed boxes and randomly replacing three plants per population with the purple variety IR1552 to represent off-types. DNA isolated from bulked leaf tissue samples of rows and columns (20 samples per population) were subjected to AFLP protocols using 32 P-labeled primers. Rows and columns involving the OFF-TYPE IR1552 showed a fingerprint pattern different (with extra bands) from the control hybrid and were considered as impure rows and columns. Plants located on hills where the impure rows and columns intersect were considered as suspected off-types from which percent seed purity can be directly derived. This strategy maintains the statistical significance of having 100 plants per grow-out performed using the conventional visual observation method and increases the efficiency of DNA sampling by reducing the number of samples from 100 (individual DNA sampling) to 20 (bulked sampling while still able to exactly identify which plants are off-types. The strategy, which was originally designed for hybrid rice seed production, can also be used for pure line varieties of rice and other crops
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