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Mini-scale Genomic DNA Extraction from Cotton

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Abstract

Large amounts of polyphenolics in cotton leaves make it difficult to obtain high-quality genomic DNA during extraction. A procedure to isolate nuclear DNA from local cotton leaves (gossypium hirsutum, MNH93, CIM443, FH672) was therefore developed. It consists of rapid isolation of stable nuclei, which hinders covalent interactions with phenolics, followed by DNA extraction. The yield and quality of the resulting DNA is satisfactory and the protocol can be scaled up or down according to sample size. It is suitable for PCR and the restriction enzyme digestion needed for Southern and RFLP analysis.

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References

  1. Buchanan-Wollaston V, Ainsworth C: Leaf senescence in Brassica napus: cloning of senescence related genes by subtractive hybridization. Plant Mol Biol 33: 821–834 (1997).

    Google Scholar 

  2. Burnell JN: Sulfate assimilation in C4 plants. Plant Physiol 75: 873–875 (1984).

    Google Scholar 

  3. Cacco G, Saccomani M, Ferrari G: Development of sulfate uptake capacity and ATP-sulfurylase activity during root elongation in maize. Plant Physiol 60: 582–584 (1977).

    Google Scholar 

  4. Cherest H, Kerjan P, Surdin-Kerjan Y: The Saccharomyces cerevisiae MET3 gene: Nucleotide sequence and relationship of the 50 non-coding region to that of MET 25. Mol Gen Genet 210: 307–313 (1987).

    Google Scholar 

  5. Clarkson DT, Luttge U: Mineral nutrition: Inducible and repressible nutrient transport systems. Prog Bot 52: 61–83 (1991).

    Google Scholar 

  6. Clarkson DT, Hawkesford MJ, Davidian JC: Membrane and long-distance transport of sulfate. In: de Kok LJ, Stulen I, Rennenberg H, Brunold C, Rauser WE (eds) Sulfur Nutrition and Sulfur Assimilation in Higher Plants, pp. 3–19. SPB Academic Publishing, The Hague, Netherlands (1993).

    Google Scholar 

  7. de Kok LJ, de Kan PJL, Tanczos OG, Kuiper PJC: Sulphate induced accumulation of glutathione and frost tolerance of spinach leaf tissue. Physiol Plant 53: 435–438 (1981).

    Google Scholar 

  8. Don RH, Cox PT, Wainwright BJ, Baker K, Mattick JS: ‘Touchdown’ PCR to circumvent spurious priming during gene amplification. Nucl Acids Res 19: 4008 (1991).

    Google Scholar 

  9. Felsenstein J: PHILYP: Phylogeny Inference Package, Version 3.5c (1993).

  10. Gutierrez-Marcos JF, Roberts MA, Campbell EI, Wray JL: Threemembers of a novel small gene-family from Arabidopsis thaliana able to complement functionally an Escherichia coli mutant defective in PAPS reductase activity encode proteins with a thioredoxin-like domain and ‘APS reductase’ activity. Proc Natl Acad Sci USA 93: 13377–13382 (1996).

    Google Scholar 

  11. Haller E, Suter M, Brunold C: Regulation of ATP sulfurylase and adenosine 50-phosphosulfate transferase by the sulfur and the nitrogen source in hetrotrophic cell suspension cultures of Paul's Scarlet rose. J Plant Physiol 125: 275–283 (1986).

    Google Scholar 

  12. Hell R, Bergmann L:-Glutamylcysteine synthetase in higher plants: catalytic properties and subcellular localization. Planta 180: 603–612 (1990).

    Google Scholar 

  13. Hell R: Molecular physiology of plant sulfur metabolism. Planta 202: 138–148 (1997).

    Google Scholar 

  14. Herschbach C, Rennenberg H: Influence of glutathione (GSH) on net uptake of sulphate and sulphate transport in tobacco plants. J Exp Bot 45: 1069–1076 (1994).

    Google Scholar 

  15. Jolivet P: Elemental sulfur in agriculture. In: de Kok LJ, Stulen I, Rennenberg H, Brunold C, Rauser WE (eds) Sulfur Nutrition and Sulfur Assimilation in Higher Plants, pp. 193–206. SPB Academic Publishing, The Hague, Netherlands (1993).

    Google Scholar 

  16. Klonus D, Hofgen R, Willmitzer L, Riesmeier JW: Isolation and characterization of two cDNA clones encoding ATP-sulfurylases from potato by complementation of a yeast mutant. Plant J 6: 105–112 (1994).

    Google Scholar 

  17. Kouchi H, Hata, S: Isolation and characterization of novel nodulin cDNAs representing genes expressed at early stages of soybean nodule development. Mol Gen Genet 238: 106–119 (1993).

    Google Scholar 

  18. Lappartient AG, Touraine B: Demand-driven control of root ATP sulfurylase activity and SO4 2_uptake in intact canola. Plant Physiol 111: 147–157 (1996).

    Google Scholar 

  19. Leustek T, Murillo M, Cervantes M: Cloning of a cDNA encoding ATP sulfurylase from Arabidopsis thaliana by functional expression in Saccharomyces cerevisiae. Plant Physiol 105: 897–902 (1994).

    Google Scholar 

  20. Leustek T: Molecular genetics of sulfate assimilation in plants. Physiol Plant 97: 411–419 (1996).

    Google Scholar 

  21. Logan HM, Cathala N, Grignon C, Davidian JC: Cloning of a cDNA encoded by a member of the Arabidopsis thaliana ATP sulfurylase multigene family. J Biol Chem 271: 12227–12233 (1996).

    Google Scholar 

  22. Maggioni A, Renosto F: Cysteine and methionine regulation of sulphate uptake in potato tuber discs (Solanum tuberosum). Physiol Plant 39: 143–147 (1977).

    Google Scholar 

  23. Murillo M, Leustek T: Adenosine-50-triphosphate-sulfurylase from Arabidopsis thaliana and Escherichia coli are functionally equivalent but structurally and kinetically divergent: nucleotide sequence of two adenosine-50-triphosphatesulfurylase cDNAs from Arabidopsis thaliana and analysis of a recombinant enzyme. Arch Biochem Biophys 323: 195–204 (1995).

    Google Scholar 

  24. Nakai K, Kanehisa M: A knowledge base for predicting protein localization sites in eukaryotic cells. Genomics 14: 897–911 (1992).

    Google Scholar 

  25. Ng A, Blomstedt CK, Gianello R, Hamill JD, Neale AD, Gaff DF: Isolation and characterization of a lowly expressed cDNA from the resurrection grass Sporobolus stapfianus with homology to eukaryote sulfate transporter proteins. Plant Physiol 111: 651 (1996).

    Google Scholar 

  26. Petrucco S, Bolchi A, Foroni C, Percudani R, Rossi GL, Ottonello S: A maize gene encoding an NADPH binding enzyme highly homologous to isoflavone reductases is activated in response to sulfur starvation. Plant Cell 8: 69–80 (1996).

    Google Scholar 

  27. Saccomani M, Cacco G, Ferrari G: Efficiency of the first steps of sulfate utilization by maize hybrids in relation to their productivity. Physiol Plant 53: 101–104 (1981).

    Google Scholar 

  28. Sambrook J, Fritsch EF, Maniatis T: Molecular Cloning: A Laboratory Manual, 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY (1989).

    Google Scholar 

  29. Sanger F, Nicklen S, Coulson AR: DNA sequencing with chain terminating inhibitors. Proc Natl Acad Sci USA 74: 5463–5467 (1977).

    Google Scholar 

  30. Schmidt A: Regulation of sulfur metabolism in plants. Prog Bot 48: 133–150 (1986).

    Google Scholar 

  31. Setya A, Murillo M, Leustek T: Sulfate reduction in higher plants: molecular evidence for a novel 50-adenylsulfate reductase. Proc Natl Acad Sci USA 93: 13383–13388 (1996).

    Google Scholar 

  32. Smith FW, Hawkesford MJ, Prosser IM, Clarkson, DT: Isolation of a cDNA from Saccharomyces cerevisiae that encodes a high affinity sulphate transporter at the plasma membrane. Mol Gen Genet 247: 709–715 (1995).

    Google Scholar 

  33. Smith FW, Ealing PM, Hawkesford MJ, Clarkson DT: Plant members of a family of sulfate transporters reveal functional subtypes. Proc Natl Acad Sci USA 92: 9373–9377 (1995).

    Google Scholar 

  34. Smith FW, Hawkesford MJ, Ealing PM, Clarkson DT, van den Berg PJ, Belcher AR, Warrilow AGS: Regulation of expression of a cDNA from barley roots encoding a high affinity sulphate transporter. Plant J 12: 875–884 (1997).

    Google Scholar 

  35. Sorbo B: Sulfate: turbidimetric and nephelometric methods. Methods Enzymology 143: 3–6 (1987).

    Google Scholar 

  36. Stulen I, de Kok LJ: Whole plant regulation of sulfurmetabolism: a theoretical approach and comparison with current ideas on regulation of nitrogen metabolism. In: de Kok LJ, Stulen I, Rennenberg H, Brunold C, Rauser WE (eds) Sulfur Nutrition and Sulfur Assimilation in Higher Plants, pp. 77–91. SPB Academic Publishing, The Hague, Netherlands (1993).

    Google Scholar 

  37. Takahashi H, Sasakura N, Noji M, Saito K: Isolation and characterization of a cDNA encoding a sulfate transporter from Arabidopsis thaliana. FEBS Lett 392: 95–99 (1996).

    Google Scholar 

  38. Takahashi H, Yamazaki M, Sasakura N, Watanabe A, Leustek T, Engler JD, Engler G, Van Montagu M, Saito K: Regulation of sulfur assimilation in higher plants: a sulfate transporter induced in sulfate-starved roots plays a central role in Arabidopsis thaliana. Proc Natl Acad Sci USA 94: 11102–11107 (1997).

    Google Scholar 

  39. Thompson JD, Higgins DG, Gibson TJ: CLUSTALW: Improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position specific gap penalties and weight matrix choice. Nucl Acids Res 22: 4673–4680 (1994).

    Google Scholar 

  40. Tietze F: Enzymic method for quantitative determination of nanogram amounts of total and oxidized glutathione: applications to mammalian blood and other tissues. Anal Biochem 27: 502–522 (1968).

    Google Scholar 

  41. Wilkinson JQ, Crawford NM: Identification of the Arabidopsis CHL3 gene as the nitrate reductase structural gene NIA2. Plant Cell 3: 461–471 (1991).

    Google Scholar 

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Authors and Affiliations

  1. National Centre of Excellence in Molecular Biology, University of the Punjab, Canal Bank Road, Thokar Niaz Baig, Lahore, 53700, Pakistan

    Bushra Chaudhry, Afshan Yasmin, Tayyib Husnain & S. Riazuddin

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  1. Bushra Chaudhry
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  2. Afshan Yasmin
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  3. Tayyib Husnain
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  4. S. Riazuddin
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Chaudhry, B., Yasmin, A., Husnain, T. et al. Mini-scale Genomic DNA Extraction from Cotton. Plant Molecular Biology Reporter 17, 280 (1999). https://doi.org/10.1023/A:1007629715971

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