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Over-expression of TsCBF1 gene confers improved drought tolerance in transgenic maize

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Abstract

The DREB/CBF transcription factors play important roles during low temperature, drought, and high-salt stress in higher plants. In this paper, we transferred TsCBF1 gene from a dicotyledonous halophyte Thellungiella halophila into the monocotyledonous crop maize (Zea mays L.). PCR, Southern blot, and RT-PCR analysis indicated that the TsCBF1 gene had been integrated into the genome of transgenic plants and was expressed in their progeny. After 14 days of drought stress treatments, transgenic plants showed improved drought tolerance with higher Relative Water Content (RWC), higher solute accumulation, and less cell damage compared with wild-type (WT) plants. Most importantly, they showed shorter anthesis-silking interval (ASI) and produced much higher grain yield than WT under drought stress. The results indicate that the TsCBF1 gene conferred enhanced drought tolerance to maize plants and may have utility for improving tolerance to other abiotic stresses as well.

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Abbreviations

ALS:

Acetolactate synthase

ASI:

Anthesis-silking interval

CBF:

C-repeat-binding factor

CRT:

C-repeat

DRE:

Dehydration-responsive element

DREB:

DRE binding protein

MDA:

Malondialdehyde

RWC:

Relative water content

TCA:

Trichloroacetic acid

WT:

Wild-type

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Acknowledgments

This research was supported by Hi-Tech Research and Development (863) Program of China (2007AA10Z175) and National Key Technologies R&D Program (2007BAD31B01)

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Authors and Affiliations

  1. School of Life Science, Shandong University, 27 Shanda South Road, 250100, Jinan, People’s Republic of China

    Shujuan Zhang, Ning Li, Feng Gao, Aifang Yang & Juren Zhang

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  1. Shujuan Zhang
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  2. Ning Li
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  3. Feng Gao
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  5. Juren Zhang
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Corresponding author

Correspondence to Juren Zhang.

Electronic supplementary material

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11032_2009_9385_MOESM1_ESM.tif

Supplemental Figure 1. Molecular characterization of T2 transgenic maize plants. (a) Southern blot analysis: lane1, DNA marker; lane2, DNA from WT plants; lane 3-7, DNA from independent transgenic linesL1, L2, L3, L4, 30 µg DNA was digested with EcoRI before being separated on agarose, and hybridized to a DIG-labeled TsCBF1 probe. (b) Southern blot analysis: lane1, DNA marker; lane2, DNA from WT; lane3-7: DNA from independent transgenic lines L1, L2, L3, L4, 30 µg DNA from WT and transgenic plants were digested with restriction enzyme BamHI, a DIG-labeled TsCBF1 probe was used as hybridization probe. (c) RT-PCR results: Lane1-4, independent transgenic lines L1, L2, L3 and L4 respectively; lane5, WT plants. A 380 bp actin fragment was amplified as an internal control. (d) Relative expression level of the TsCBF1 gene between transgenic maize plants in T2 and T4 generation: L1, L2, L3 and L4: T2 independent transgenic lines; P1, P2, P3 and P4: T4 independent transgenic lines from L1, L2, L3 and L4. Relative RNA transcripts were calculated by the 2−ΔΔCt method (Livak and Schmittgen 2001), with maize actin as an internal control. All experiments were repeated at least three times. (TIF 47298 kb)

11032_2009_9385_MOESM2_ESM.tif

Supplemental Figure 2. Expression changes of putative downstream genes of CBF1 under drought stress. a. Expression analysis of transgenic plants; b. Expression patterns of WT plants; c. relative expression levels calculated by the ratio of fold changes of transgenic plants/fold changes of WT. Total RNA was isolated from the leaves of 10 leaves maize plants at 0d, 1d, 3d, 5d after drought stress. Expression patterns of these genes were analyzed by real-time RT-PCR, and fold changes of RNA transcripts were calculated by the 2−ΔΔCt method (Livak and Schmittgen 2001) with maize actin as an internal control. Values are means of replicates with standard deviation. The entire experiments were repeated three times. (TIFF 7992 kb)

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Zhang, S., Li, N., Gao, F. et al. Over-expression of TsCBF1 gene confers improved drought tolerance in transgenic maize. Mol Breeding 26, 455–465 (2010). https://doi.org/10.1007/s11032-009-9385-5

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