Obtaining high-quality plant materials for experiments is challenging for many research projects.Therefore, it is of special importance to determine the best method for preserving biological macromolecules like DNA, w...Obtaining high-quality plant materials for experiments is challenging for many research projects.Therefore, it is of special importance to determine the best method for preserving biological macromolecules like DNA, which degrade over time. Although some research has demonstrated that DNA degradation has little effect on traditional molecular markers, the effects of DNA degradation on dd RADseq, a popular reduced-representation sequencing technology, have not been adequately investigated. In this study, we first chose six woody bamboo species(Bambusoideae, Poaceae) to explore appropriate methods for preserving molecular materials with two DNA extraction approaches. Then we sequenced twenty-one bamboos and examined the effects of DNA quality on data generation using the dd RAD-seq technique(Midd RAD-seq). Finally, we reconstructed phylogenies of twenty woody bamboo species. We found that the integrity of dry-powdered DNA was preserved longer than that of TE-dissolved DNA,regardless of whether the DNA was extracted by a modified CTAB protocol or DNAsecure plant kit. The dd RAD-seq data were robust, except when DNA was severely degraded. In addition, we resolved the phylogenetic positions of the sampled Phyllostachys spp. Our results suggest that dry-powdered DNA is the most appropriate preservation method for plant molecular materials. Furthermore, a moderate level of DNA degradation has little effect on reduced representation sequencing techniques represented by dd RAD-seq.展开更多
基金supported by the National Natural Science Foundation of China (Grant No. 31470322 and 31430011)
文摘Obtaining high-quality plant materials for experiments is challenging for many research projects.Therefore, it is of special importance to determine the best method for preserving biological macromolecules like DNA, which degrade over time. Although some research has demonstrated that DNA degradation has little effect on traditional molecular markers, the effects of DNA degradation on dd RADseq, a popular reduced-representation sequencing technology, have not been adequately investigated. In this study, we first chose six woody bamboo species(Bambusoideae, Poaceae) to explore appropriate methods for preserving molecular materials with two DNA extraction approaches. Then we sequenced twenty-one bamboos and examined the effects of DNA quality on data generation using the dd RAD-seq technique(Midd RAD-seq). Finally, we reconstructed phylogenies of twenty woody bamboo species. We found that the integrity of dry-powdered DNA was preserved longer than that of TE-dissolved DNA,regardless of whether the DNA was extracted by a modified CTAB protocol or DNAsecure plant kit. The dd RAD-seq data were robust, except when DNA was severely degraded. In addition, we resolved the phylogenetic positions of the sampled Phyllostachys spp. Our results suggest that dry-powdered DNA is the most appropriate preservation method for plant molecular materials. Furthermore, a moderate level of DNA degradation has little effect on reduced representation sequencing techniques represented by dd RAD-seq.
