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Korean Society for Biochemistry and Molecular Biology (생화학분자생물학회)
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Construction and Differential Screening of a cDNA Library Specific to Osmotic Stress of Haloxylon ammodendron Seedlings

  • Jiang, Xiao-Cheng (College of Life Science, Hunan Normal University) ;
  • Guo, Xin-Hong (College of Life Science, Hunan Normal University) ;
  • Pan, Xiao-Ling (Arid Ecological Environment Research Institute of Xinjiang University) ;
  • Song, Song-Quan (Xishuangbanna Tropical Botanical Garden, The Chinese Academy of Science)
  • Published : 2004.09.30
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Abstract

A subtracted cDNA library specific to osmotic stress of Haloxylon ammodendron (Mey.) Bge seedlings was constructed by suppression subtractive hybridization (SSH) and T/A cloning. SSH was performed between two groups of H. ammodendron seedlings, one was cultivated in Hoagland (H) solution as a driver and the other group was treated with osmotic stress of the Hoagland solution by the addition of 400 mM mannitol (M), as a tester. The library consisted of about 400 recombinant clones, with the average size being of 500 bp, ranging from 300 bp to 1500 bp. Using a PCR-select differential screening kit, 100 recombinant clones were randomly chosen from the subtracted cDNA library and hybridized with forward,reverse subtracted and unsubtracted probes for two rounds. As a result, 21 positive clones specific to osmotic stress were obtained and some of them were verified by Northern blot analysis. The sequencing analysis of 6 positive clones and the following homology comparison to GenBank [blastx] non-redundant databases characterized that two sequences obtained in this experiment may contribute to novel drought-related genes.

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References

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