Journal
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
Volume 101, Issue -, Pages 595-602Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.ijbiomac.2017.03.062
Keywords
Antifreeze protein; Ortholog; Thermal hysteresis
Funding
- Special Fund for Forestry-Scientific Research in the Public Interest of China [201504101]
- Fundamental Research Funds for the Central Universities grants of China [2015ZCQ-SW-01]
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Spruce can grow at an extra low temperature (LT), and is inferred with, important antifreezing gene resources. The research here identified 4 different spruce varieties, named as PicW1, PicW2, PicM and PicK. Sequence alignment showed base-substitution and deficiency mutations among them with sequence identity between 97.61% and 99.25%. Each gene was transferred into E. coli, where protein was induced by IPTG (isopropyl-beta-n-thiogalactoside). Strains cultured at -5 degrees C showed the lethal dose 50% (LD-50) between 53 h and 57 h for the transgenic strains, but 35 h for the control. Strains cultivated at 20 degrees C showed the LD-50 between 38 h and 44h for the transgenic strains, but 25 h for the control. Further, the soluble gene proteins were extracted and purified for Differential Scanning Calorimeter (DSC) test, which showed characteristic thermal hysteresis (TH) value of 0.77 degrees C (PicW1), 0.78 degrees C (PicW2), 0.72 degrees C (PicM), and 0.86 degrees C (PicK) respectively, significantly higher than the value of 0.05 degrees C of the control (BSA). Summarily, four homologous proteins showed good antifreeze property with the range from high to low as PicK > PicW2 > PicW1 > PicM. It suggested that they can be used as resources for genetic engineering of plant cold tolerance. (C) 2017 Published by Elsevier B.V.
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