DNA Fingerprint of the Current Recommended Field Crop Varieties
Nareuthai Worasathit, Summana Ngampongsai, Haiporn Kittikul and Chantana Kongnakhon
Field Crops Research Institute, Department of Agriculture, Chatuchak, Bangkok 10900, Thailand.
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Biotechnology, especially in the area of molecular genetics, is now receiving a lot more attention from people all over the world. It has been said that biotechnology is potentially the new green revolution in agriculture. One example of a current advancement made in biotechnology is the development of the cotton variety, known as Bollgard of NuCOTN 32. This has been engineered to resist the American boll-worn pest by transferring the bt-gene or cry 1 A ( c), normally present in Bacillus thruringiensis sub sp. Kurstaki into the cotton variety, Coker 315 and then DP 5415 (DPX 174).
Another potential benefit that could be derived from biotechnology is the development of DNA fingerprinting. This technique can be used in variety identification to detect the true variety and variety purity. Other potential benefits of DNA fingerprinting include gene mapping, marker-assisted selection and assessment of genetic diversity. Much emphasis has currently been given to these areas of research.
In Thailand, the Plant Variety Protection Act of 1999 was put into action in order to protect the right of, not only of the plant breeder but also the farmer. New variety that can be covered under the Act should have distinct characteristics from those of the current varieties, together with possessing the qualities of uniformity and stability. Such variety characterization can be determined from the morphology of the plant. In addition, DNA fingerprinting technology will also help variety identification to be more precise.
In order to set up a database of the DNA fingerprints of the current recommended field crop varieties, a research program was initiated in collaboration with the Faculty of Arts and Science at Kasetsart University in 1999. The technique used was that of the Random Amplified Polymorphic DNA (RAPD). The procedure consisted of sample preparation, DNA extraction, RAPD reaction (using 6 primers as follow, primer 1:5’GGTGCGGGAA-3’, Primer 2:5’-GTTTCGCTCC-3’, 3:5’-CCCGTCAGCA-3’), primer 4:5’- AAAAGCCCGT-3’, primer 5:5’AACGCGCAAC-3’, and 6:5’-CCCGTCAGCA-3’), PCR reaction and DNA pattern analysis. Two replicates were made from each variety.
The results indicated that each variety could be consistently identified using the RAPD technique. Relationships between varieties could also be analysed and shown as a dendrogram. In some crops, such as sorghum, at least 2 or more primers were required to differentiate between varieties. For groundnut and cotton, the RAPD technique was not applicable and polymorphism or variation among varieties could not be observed even though several primers had been used. The limitations to the use of this technique may be due to the close genetic relationships among varieties and the seed purity as well as heterogeneity of those varieties. Further study is required to precisely identify these crop varieties using more specific techniques. Figures below show the DNA profile of some field crops used in the study.
DNA