Assessment of Recovery Medium for Production of hCTLA4Ig after Cryopreservation in Transgenic Rice Cells

다국어 초록 (Multilingual Abstract)

A reproducible method for cryopreservation of transgenic rice cells (Oryza sativa L. cv. Dongjin) producing recombinant human cytotoxic T lymphocyte-associated antigen 4-immunoglobulin (hCTLA4Ig) has been established.
Here, we assessed recovery media and investigated recombinant protein homogeneity after long-term preservation.
For recovery of cryopreserved transgenic rice cells, AA medium was suitable in terms of both morphology and production of hCTLA4Ig. There were no differences in cell growth, sugar consumption, and hCTLA4Ig production between non-cryopreserved and cryopreserved cells for up to 1 month. hCTLA4Ig produced from cryopreserved cells was identical that of hCTLA4Ig from non-cryopreserved cells, as determined by analysis of its molecular weight and isoforms. For long-term preservation, cell viability was stably maintained at 61% for 26 months. In conclusion, these results demonstrate the possibility for reproducible cryocell-banking of transgenic rice cells without changes in the characteristics of cells and target proteins.

참고문헌 (Reference)

1 Zhu, L., "Versatile characterization of glycosylation modification in CTLA4-Ig fusion proteins by liquid chromatography-mass spectrometry" 6 : 1474-1485, 2014

2 Jain, M., "Validation of housekeeping genes as internal control for studying gene expression in rice by quantitative real-time PCR" 345 : 646-651, 2006

3 Wesley-Smith J., "The influence of water content, cooling and warming rate upon survival of embryonic axes of Poncirus trifoliata (L.)" 25 : 129-138, 2004

4 Rabilloud, T., "Solubilization of proteins for electrophoretic analyses" 17 : 813-829, 1996

5 Drakakaki, G., "Protein secretion: How many secretory routes does a plant cell have?" 203-204 : 74-78, 2013

6 Faye, L., "Protein modifications in the plant secretory pathway: current status and practical implications in molecular pharming" 23 : 1770-1778, 2005

7 Jung, J. -W., "Production and characterization of recombinant human acid ${\alpha}$-glucosidase in transgenic rice cell suspension culture" 226 : 44-53, 2016

8 Strasser, R., "Plant protein glycosylation" 26 : 926-939, 2016

9 James, E., "Loss and recovery of protein productivity in genetically modified plant cell lines" 25 : 723-727, 2006

10 Whitmer, S., "Long-term instability of alkaloid production by stably transformed cell lines of Catharanthus roseus" 74 : 73-80, 2003

11 Tekoah, Y., "Large-scale production of pharmaceutical proteins in plant cell culture-the protalix experience" 13 : 1199-1208, 2015

12 Mustafa, N. R., "Initiation, growth and cryopreservation of plant cell suspension cultures" 6 : 715-742, 2011

13 Kaczmarczyk, A., "Influence of alternating temperature preculture on cyropreservation results for potato shoot tips" 27 : 1551-1558, 2008

14 Kuo, Y. -C., "Improving pharmaceutical protein production in Oryza sativa" 14 : 8719-8739, 2013

15 Kang, S. -H., "Glycan structure and serum half-life of recombinant CTLA4Ig, an immunosuppressive agent, expressed in suspensioncultured rice cells with coexpression of human ${\beta}1$,4-galactosyltransferase and human CTLA4Ig" 32 : 161-172, 2015

16 Hayashi, T., "Effects of amino acid medium on cell aggregation in suspension-cultured rice cells" 58 : 256-260, 1994

17 Dahyun Yu, "Effects of Culture Media on hCTLA4Ig Production and Protein Expression Patterns in Transgenic Rice Cell Suspension Cultures" 한국생물공학회 13 (13): 424-430, 2008

18 Cho, J. -S., "Cryopreservation of transgenic rice suspension cells producing recombinant hCTLA4Ig" 73 : 1470-1476, 2007

19 Lullau, E., "Comparison of batch and perfusion culture in combination with pilot-scale expanded bed purification for the production of soluble recombinant ${\beta}$-secretase" 19 : 37-44, 2003

20 Sethuraman, N., "Challenges in therapeutic glycoprotein production" 17 : 341-346, 2006

21 Rosales-Mendoza, S., "Carrot cells: A Pioneering platform for biopharmaceuticals production" 57 : 219-232, 2015

22 Gomord, V., "Biopharmaceutical production in plants: problems, solutions and opportunities" 23 : 559-565, 2005

23 Kwon, J. -Y., "Assessment of long-term cryopreservation for production of hCTLA4Ig in transgenic rice cell suspension cultures" 53 : 216-222, 2013

24 Livak, K. J., "Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method" 24 : 402-408, 2001

25 Boisson, A. -M., "A simple and efficient method for the long-term preservation of plant cell suspension cultures" 8 : 4-, 2012