A Simple and Efficient Method for High Quality DNA Extraction from Sweet Sorghum [Sorghum bicolor (L.) Moench] (original) (raw)
Related papers
IJCS, 2020
Extraction of genomic DNA from plant leaf tissue is a basic requirement for molecular biology experiment as well as in plant breeding for crop improvement. The purity of DNA is affected by varying level of polyphenols, polysaccharides and secondary metabolites with are associated with DNA at the time of extraction. A variety of DNA extraction methods and kits are available, however they are costly, low yield and time consuming. Here, we describe a simple and efficient method of DNA extraction from sorghum leaf. The present study is based on conventional cetyl trimethyl ammonium bromide (CTAB) method with four major modifications to isolate genomic DNA from seven different accessions of sorghum and the yield of DNA was varied from1550.2 ngµl-1 to 1910.5ngµl-1 in 100 µl final volume of TE buffer. The purity of DNA sample was found to be varied from 1.53 to 2.16 based on the absorbance at A260 and A280 ratio. This method is very simple as it not requires liquid nitrogen or magnetic beads to grind the leaf sample which is mostly unavailable to some undergraduate laboratories.
A high throughput DNA extraction method with high yield and quality
Plant Methods, 2012
Background: Preparation of large quantity and high quality genomic DNA from a large number of plant samples is a major bottleneck for most genetic and genomic analyses, such as, genetic mapping, TILLING (Targeting Induced Local Lesion IN Genome), and next-generation sequencing directly from sheared genomic DNA. A variety of DNA preparation methods and commercial kits are available. However, they are either low throughput, low yield, or costly. Here, we describe a method for high throughput genomic DNA isolation from sorghum [Sorghum bicolor (L.) Moench] leaves and dry seeds with high yield, high quality, and affordable cost. Results: We developed a high throughput DNA isolation method by combining a high yield CTAB extraction method with an improved cleanup procedure based on MagAttract kit. The method yielded large quantity and high quality DNA from both lyophilized sorghum leaves and dry seeds. The DNA yield was improved by nearly 30 fold with 4 times less consumption of MagAttract beads. The method can also be used in other plant species, including cotton leaves and pine needles. Conclusion: A high throughput system for DNA extraction from sorghum leaves and seeds was developed and validated. The main advantages of the method are low cost, high yield, high quality, and high throughput. One person can process two 96-well plates in a working day at a cost of $0.10 per sample of magnetic beads plus other consumables that other methods will also need.
Whole‐genomic DNA amplifications from individually isolated sweet sorghum microspores
Applications in Plant Sciences, 2022
PremiseSorghum is a multi‐use crop, the efficient breeding of which requires the development of new genetic tools. One such tool could be the genetic assessment of free microspores, which are released just after the tetrad stage of pollen development. Microspores are ideal for DNA isolation as they have underdeveloped cell walls and can be readily lysed as natural protoplasts.MethodsFour cultivars of Sorghum bicolor (ʻAchi Turi’, ʻDale’, ʻLocal’, and ʻTopper 76‐6’) were grown in a greenhouse until flowering (7.7–11.5 cm flag leaf internode length), after which 30 immature microspores were isolated from each line. Plant height, time to flowering, boot radius, and spikelet maturation were recorded for each cultivar. The exine development of the microspores was observed under an inverted Nikon microscope, and those with underdeveloped exine were subjected to whole‐genome amplification and sequencing.ResultsMicrospores in the early uninucleate to early binucleate stages had underdeveloped exine, and were therefore ideal for DNA extraction. High‐quality DNA was obtained from these single‐cell gametophytes. The average DNA concentration was 2902 ng/µL, with fragment sizes comparable to those obtained from leaf tissue extractions.DiscussionHarvesting panicles with immature microspores means the entire gametic population is accessible for DNA analyses. This is the first amplification of whole‐genome DNA fragments from sorghum single‐cell microspores isolated during gametogenesis.
Extraction of genomic DNA from plant tissues
Background: Preparation of large quantity and high quality genomic DNA from a large number of plant samples is a major bottleneck for most genetic and genomic analyses, such as, genetic mapping, TILLING (Targeting Induced Local Lesion IN Genome), and next-generation sequencing directly from sheared genomic DNA. A variety of DNA preparation methods and commercial kits are available. However, they are either low throughput, low yield, or costly. Here, we describe a method for high throughput genomic DNA isolation from sorghum [Sorghum bicolor (L.) Moench] leaves and dry seeds with high yield, high quality, and affordable cost. Results: We developed a high throughput DNA isolation method by combining a high yield CTAB extraction method with an improved cleanup procedure based on MagAttract kit. The method yielded large quantity and high quality DNA from both lyophilized sorghum leaves and dry seeds. The DNA yield was improved by nearly 30 fold with 4 times less consumption of MagAttract beads. The method can also be used in other plant species, including cotton leaves and pine needles. Conclusion: A high throughput system for DNA extraction from sorghum leaves and seeds was developed and validated. The main advantages of the method are low cost, high yield, high quality, and high throughput. One person can process two 96-well plates in a working day at a cost of $0.10 per sample of magnetic beads plus other consumables that other methods will also need.
High throughput BAC DNA isolation for physical map construction of sorghum (Sorghum bicolor)
1998
With the aim of constructing a physical map of sorghum, we developed a rapid, high throughput approach for isolating BAC DNA suitable for restriction endonuclease digestion fingerprinting, PCR-based STS-content mapping, and BAC-end sequencing. The system utilizes a programmable 96 channel liquid handling system and associated accessories that permit bacterial cultivation and DNA isolation in 96-well plate format. This protocol details culture conditions that optimize bacterial growth in deep-well plates and criteria for BAC DNA isolation to obtain high yields of quality BAC DNA. The system is robust, accurate, and relatively cost-effective. The BAC DNA isolation system has been tested during efforts to construct a physical map of sorghum.
Hindawi Publishing CorporationReview Article Genomics of Sorghum
Sorghum (Sorghum bicolor (L.) Moench) is a subject of plant genomics research based on its importance as one of the world's leading cereal crops, a biofuels crop of high and growing importance, a progenitor of one of the world's most noxious weeds, and a botanical model for many tropical grasses with complex genomes. A rich history of genome analysis, culminating in the recent complete sequencing of the genome of a leading inbred, provides a foundation for invigorating progress toward relating sorghum genes to their functions. Further characterization of the genomes other than Saccharinae cereals may shed light on mechanisms, levels, and patterns of evolution of genome size and structure, laying the foundation for further study of sugarcane and other economically important members of the group.
“ABBAS” DNA Extraction Method from plant
2013
DNA extraction is still problematic in a variety of plants because of the presence of secondary metabolites that interfere with DNA isolation procedures and downstream applications such as DNA restriction, amplification, and cloning. Here we describe a modified procedure based on the polyethylene glycol (PEG-6000) method to isolate DNA from tissues containing high levels of polysaccharides and phenolics. The procedure is applicable for sugarcane, olive, wheat, sorghum, barley, gram, sea buckthorn, amla, autumn olive and related species form leaves and fruits. This modified method contains PEG (1%), 4M NaCl, 0.5M EDTA, 1MTris HCl and SDS (10%) and washed, also reduced the centrifugation times during the separation and precipitation of the DNA. The method has solved the problems of DNA degradation, contamination, and low yield due to binding and/or coprecipitation with starch and polysaccharides. The isolated DNA proved amenable to PCR amplification and restriction digestion. The tech...
A General Overview of Sweet Sorghum Genomics
Biotechnological Applications of Biomass
Sorghum is one of the main cereal crops, its consumption is large, since it provides grain, fiber and biofuel. Likewise, its genome, with only 10 diploid chromosomes, makes it an attractive model for research and genetic improvement. Sorghum is the most studied C4 plant of its genus; several lines have been developed under three main characteristics: grain, forage and sugar biomass. Compared to other crops, sweet sorghum possesses high levels of highly fermentable sugars in the stem. Also, it has the ability of producing high production yields in marginal lands. These characteristics make it and attractive crop for the generation of biofuels. Molecular markers associated to several resistances and tolerances to biotic and abiotic factors have been described in literature. These allow the development of high-density linkage maps, which, along with the rising availability of sorghum genomes, will accelerate the identification of markers and the integration of the complete genome seque...
Sorghum genetic, genomic, and breeding resources
Planta, 2021
Main conclusion Sorghum research has entered an exciting and fruitful era due to the genetic, genomic, and breeding resources that are now available to researchers and plant breeders. Abstract As the world faces the challenges of a rising population and a changing global climate, new agricultural solutions will need to be developed to address the food and fiber needs of the future. To that end, sorghum will be an invaluable crop species as it is a stress-resistant C4 plant that is well adapted for semi-arid and arid regions. Sorghum has already remained as a staple food crop in many parts of Africa and Asia and is critically important for animal feed and niche culinary applications in other regions, such as the United States. In addition, sorghum has begun to be developed into a promising feedstock for forage and bioenergy production. Due to this increasing demand for sorghum and its potential to address these needs, the continuous development of powerful community resources is requ...