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Development of highly regenerable callus lines and Agrobacterium-mediated transformation of Chinese lawngrass (Zoysia sinica Hance) with a cold inducible transcription factor, CBF1

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Abstract

Chinese lawngrass (Zoysia sinica Hance) is a warm-season turfgrass, which is widely used for golf courses, athletic fields and home lawns. In order to improve Chinese lawngrass with biotechnology, an efficient plant regeneration system was established. Five highly regenerable callus lines have been developed by addition of 6-benzylaminopurine (6-BA) and adjusting concentration of 2,4-dichlorophenoxyacetic acid (2,4-D) in culture medium after multiple subculture, of which callus line Zh44 was used for Agrobacterium-mediated transformation. The gene, encoding C-repeat/dehydration-responsive element binding factor 1 (CBF1/DREB1b) from Arabidopsis driven by cauliflower mosaic virus (CaMV) 35S promoter, was introduced into Chinese lawngrass under selection using the bar gene with glufosinate. Three independent transgenic lines were successfully obtained using a three-step selection/regeneration procedure. The transgenic plants were confirmed by polymerase chain reaction (PCR), southern blot and reverse transcriptase polymerase chain reaction (RT-PCR) analyses. Two transgenic lines exhibited growth retardation and decreased tillering. The transgenic lines have a significantly stronger chilling tolerance than that of wild type plants, measured by ion leakage, total chlorophyll content and yellowed leaf rate in chilling conditions.

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Abbreviations

AS:

acetosyringone

6-BA:

benzylaminopurine

CBF1 :

C-repeat/dehydration-responsive element binding factor 1

2,4-D:

2,4-dichlorophenoxyacetic acid

MS:

Murashige and Skoog medium

NAA:

naphthaleneacetic acid

PCR:

polymerase chain reaction

RT-PCR:

reverse transcriptase polymerase chain reaction

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Acknowledgments

This work was supported by the State ‘863’ High-Tech Program (Grant No. 2002AA241061) and in part by National Natural Science Foundation of China (Grant No. 30370856). We are also grateful to Dr. Zhen-bo Tan for providing Agrobacterium strain and binary vector pBPCBF1bar for the study.

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Authors and Affiliations

  1. Bei**g Agro-Biotechnology Research Center, Bei**g Academy of Agricultural and Forestry Sciences, 100089, Bei**g, P.R. China

    Rui-fen Li, Jian-hua Wei, Hong-zhi Wang & Jie He

  2. Research Institute of Forestry, Chinese Academy of Forestry, 100091, Bei**g, P.R. China

    Zhen-yuan Sun

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  1. Rui-fen Li
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  2. Jian-hua Wei
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  3. Hong-zhi Wang
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  4. Jie He
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  5. Zhen-yuan Sun
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Correspondence to Zhen-yuan Sun.

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Li, Rf., Wei, Jh., Wang, Hz. et al. Development of highly regenerable callus lines and Agrobacterium-mediated transformation of Chinese lawngrass (Zoysia sinica Hance) with a cold inducible transcription factor, CBF1 . Plant Cell Tiss Organ Cult 85, 297–305 (2006). https://doi.org/10.1007/s11240-006-9080-8

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  • DOI: https://doi.org/10.1007/s11240-006-9080-8

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