Chinese Bulletin of Botany ›› 2022, Vol. 57 ›› Issue (3): 340-349.DOI: 10.11983/CBB21206

• TECHNIQUES AND METHODS • Previous Articles     Next Articles

An Efficient Protoplast Transient Expression System in Camellia sinensis var. sinensis cv. ‘Tieguanyin’

Yuqin Zhang1,2, Jiacheng Wu1,2, Meng He2, Renyi Liu3, Xiaoyue Zhu2,*()   

  1. 1College of Horticulture, Fujian Agriculture and Forestry University, Fuzhou 350002, China
    2FAFU-UCR Joint Center for Horticultural Plant Biology and Metabolomics, Fujian Agriculture and Forestry University, Fuzhou 350002, China
    3Center for Agroforestry Mega Data Science, Fujian Agriculture and Forestry University, Fuzhou 350002, China
  • Received:2021-11-24 Accepted:2022-02-07 Online:2022-05-01 Published:2022-05-18
  • Contact: Xiaoyue Zhu

Abstract: Recent advances in the genomic sequencing of tea plants have laid the foundation for tea research at the molecular and gene levels. However, the transgenic technologies are immature and the life cycles are long for tea trees, it is still difficult to conduct functional analyses of tea genes. This study used young leaves of Camellia sinensis var. sinensis cv. ‘Tieguanyin’, established a useful formula by testing multiple concentration combinations of cellulase, pectinase, macerozyme and mannitol. By evaluating the quantity, viability and debris of resulted protoplast, we successfully established a highly efficient mesophyll protoplast isolation and PEG-mediated transient expression system in Tieguanyin seedling leaves, with a transformation rate reaching 56.25%. Using this system, the subcellular localization of two pivotal enzymes in the theanine metabolism pathway (the theanine synthetase (TSI) and the glutamine synthetase (GSII-1.1)) were explored. Results show that, these two enzymes are both localized in the cytosol of the Tieguanyin protoplasts. Together, the establishment of this tea mesophyll protoplast extraction and transient expression system would lay a technological foundation for studying the function of tea genome.

Key words: Tieguanyin, theanine, theanine synthetase, glutamine synthetase, protoplast transfection