Cloning of Glutathione Reductase 1 in Eucalyptus grandis×E. ophylla and the Expression Pattern in Chilling Resistance

doi:10.3969/j.issn.1000-2561.2019.03.012

Abstract

Abstract:

Glutathione reductase (GR) catalyzes the conversion of oxidized glutathione to reduced glutathione (GSH). EuGR1 (GenBank Accession Number: KU904639) is located in the Eucalyptus grandis×E.ophylla cytoplasm containing 1485 bp nucleotides and encoding 495 amino acids. A prokaryotic expression vector of pET-EuGR1 was constructed and showed high GR activity in E. coli BL21. Quantitative PCR was used to analyze the spatiotemporal expression of EuGR1 in eucalyptus. The results showed that the expression of EuGR1 was the highest in the young leaves of eucalyptus seedlings and decreased with the maturity of leaves. In the tissue culture of eucalyptus the expression of EuGR1 was the highest in the stem, followed by the leaf, and the expression in the root was the lowest. The expression of EuGR1 gene in the cold-resistant transgenic eucalyptus was studied under low temperature stress. The result showed that plants with strong cold tolerance under normal temperature and low temperature stress had higher expression of EuGR1 (such as P40, P41, P52) than the plants with weak cold tolerance (such as P36, F44, F76), indicating that the expression level of EuGR1 in eucalyptus was associated with cold resistance. However, the expression of EuGR1 kept decreasing in the tested plants at low temperature treatment after 36 h suggesting that cytoplasmic EuGR1 might play an important role in the early stage of low temperature stress tolerance in Eucalyptus.

Key words: Eucalyptus grandis×E.ophylla;, glutathione reductase, clone, low temperature stress

ZHAO Yanling,HAN Yao. Cloning of Glutathione Reductase 1 in Eucalyptus grandis×E. ophylla and the Expression Pattern in Chilling Resistance[J]. Chinese Journal of Tropical Crops, 2019, 40(3): 499-503.

Figures/Tables 7

Fig. 1 WOLF-PROST subcellular localization result

Fig. 2 Enzyme digestion of pET-EuGR1 by Hind ⅢM: DNA Marker; 1: pET-EuGR1; 2: pET-32a(+)

Fig. 3 The SDS-PAGE of pET-EuGR1 expression in E. coli BL21 M: Protein marker; 1: E. coil BL21(with 1.0 mmol/L IPTG); 2: E. coil BL21; 3: pET-EuGR1 (with 1.0 mmol/L IPTG); 4: pET-EuGR1; 5: pET-32a(+)(with 1.0 mmol/L IPTG); pET-EuGR1

Fig. 4 The GR enzyme activity in E. coil BL21 with pET-EuGR1 BL21: E. coil BL2, PET: E. coil BL21 containing pET-32a, GR: E. coil BL21 containing pET-EuGR1.

Fig. 5 Relative quantitive ananlysis results of EuGR1 in Eucalyptus tissue culture seedling

Fig. 6 Relative quantitive ananlysis results of EuGR1 in Eucalyptus leaf

Fig. 7 Relative quantitive ananlysis of EuGR1 in Eucalyptus seedlings at low temperature stress

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