Development of DNA Marker for Genetic Purity Testing of Marigold Hybrid Seeds
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Abstract
Grow-Out Test (GOT) is a traditional method used to determine the genetic purity of hybrid marigold seeds. The disadvantages of these method are time-consuming, need high labor cost and huge space to maintain the seedling plants up to mature stage. In addition, evaluation based on phenotypic traits is difficult and often inaccurate, especially for closely-related genetic varieties. The research aimed to develop a highly reproducible DNA marker for genetic purity testing of hybrid seeds and differentiating the hybrid from its parental lines. The novel marker (S12+GCAG) was derived using RAPD-S12 primer, which gave an unique band size of 1,108 bp, thus distinguishing between the hybrid and its parent plants. The DNA fragment containing the marker was cloned into the pTA2 plasmid vector and sequenced. Primers were designed based on core sequence similar to S12 and adding 2, 4, 8 and 14 bases, respectively at 3’. Among the 4 designed primers, only S12 with 4 bases (+GCAG) added could be distinguished from the parental line. The marker was subjected to genetic purity testing of hybrid seeds, which artificially added with contaminated seeds. The results showed that out of 100 seeds, there were 93 true hybrid seeds, 3 contaminated seeds and 4 self-pollinated seeds. This developed primer was more stable than the RAPD DNA marker.
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King Mongkut's Agricultural Journal
References
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