Qu Junge
(Marine Bioproducts Engineering Group, Dalian Institute of Chemical Physics, Chinese Academy of Sciences)
,
Zhang Wei
(Marine Bioproducts Engineering Group, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, CRC for Bioproducts and Department of Medical Biotechnology, School of Medicine, Flinders University)
,
Yu Xingju
(Marine Bioproducts Engineering Group, Dalian Institute of Chemical Physics, Chinese Academy of Sciences)
,
Jin Meifang
(Marine Bioproducts Engineering Group, Dalian Institute of Chemical Physics, Chinese Academy of Sciences)
The inherent instability of metabolite production in plant cell culture-based bioprocessing is a major problem hindering its commercialization. To understand the extent and causes of this instability, this study was aimed at understanding the variability of anthocyanin accumulation during long-term ...
The inherent instability of metabolite production in plant cell culture-based bioprocessing is a major problem hindering its commercialization. To understand the extent and causes of this instability, this study was aimed at understanding the variability of anthocyanin accumulation during long-term subcultures, as well as within subculture batches, in Vitis vinifera cell cultures. Therefore, four cell line suspensions of Vitis vinifera L. var. Gamay Freaux, A, B, C and D, originated from the same callus by cell-aggregate cloning, were established with starting anthocyanin contents of $2.73\;\pm\;0.15,\;1.45\;\pm\;0.04,\;0.7\;\pm\;0.024\;and\;0.27\;\pm\;0.04$CV (Color Value)/g-FCW (fresh cell weight), respectively. During weekly subculturing of 33 batches over 8 months, the anthocyanin biosynthetic capacity was gradually lost at various rates, for all four cell lines, regardless of the significant difference in the starting anthocyanin content. Contrary to this general trend, a significant fluctuation in the anthocyanin content was observed, but with an irregular cyclic pattern. The variabilities in the anthocyanin content between the subcultures for the 33 batches, as represented by the variation coefficient (VC), were 58, 57, 54, and $84\%$ for V. vinifera cell lines A, B, C and D, respectively. Within one subculture, the VCs from 12 replicate flasks for each of 12 independent subcultures were averaged, and found to be $9.7\%$, ranging from 4 to $17\%$. High- and low-producing cell lines, VV05 and VV06, with 1.8-fold differences in their basal anthocyanin contents, exhibited different inducibilities to L-phenylalanine feeding, methyl jasmonate and light irradiation. The low-producing cell line showed greater potential in enhanced the anthocyanin production.
The inherent instability of metabolite production in plant cell culture-based bioprocessing is a major problem hindering its commercialization. To understand the extent and causes of this instability, this study was aimed at understanding the variability of anthocyanin accumulation during long-term subcultures, as well as within subculture batches, in Vitis vinifera cell cultures. Therefore, four cell line suspensions of Vitis vinifera L. var. Gamay Freaux, A, B, C and D, originated from the same callus by cell-aggregate cloning, were established with starting anthocyanin contents of $2.73\;\pm\;0.15,\;1.45\;\pm\;0.04,\;0.7\;\pm\;0.024\;and\;0.27\;\pm\;0.04$CV (Color Value)/g-FCW (fresh cell weight), respectively. During weekly subculturing of 33 batches over 8 months, the anthocyanin biosynthetic capacity was gradually lost at various rates, for all four cell lines, regardless of the significant difference in the starting anthocyanin content. Contrary to this general trend, a significant fluctuation in the anthocyanin content was observed, but with an irregular cyclic pattern. The variabilities in the anthocyanin content between the subcultures for the 33 batches, as represented by the variation coefficient (VC), were 58, 57, 54, and $84\%$ for V. vinifera cell lines A, B, C and D, respectively. Within one subculture, the VCs from 12 replicate flasks for each of 12 independent subcultures were averaged, and found to be $9.7\%$, ranging from 4 to $17\%$. High- and low-producing cell lines, VV05 and VV06, with 1.8-fold differences in their basal anthocyanin contents, exhibited different inducibilities to L-phenylalanine feeding, methyl jasmonate and light irradiation. The low-producing cell line showed greater potential in enhanced the anthocyanin production.
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제안 방법
For the data presented in this paper, cell line A was sub- cultured in suspension for 50 generations, and cell lines B, C and D in suspensions for 60 generations. At the time of each subsequent subculture, triplicate samples were taken, and their anthocyanin contents analyzed. The long-term subculture experiment was carried out for 33 batches over 231 days.
The experiments were carried out in 250-mL Erlenmeyer flasks containing 50 mL B5 medium supplemented with 30 g/L sucrose, 250 mg/L casein hydrolysate, 0.1 mg/L a-naphthaleneacetic acid and 0.2 mg/L kinetin, with an inoculum of 5.0 g of wet cells. For each of the four cell lines: A, B, C and D, 12 flasks were inoculated from the same parental culture and cultivated under the same subculture conditions detailed above.
vinifera. These experiments were carried out randomly during the long-term subcultures with a broad range of anthocyanin contents (from 1.33 to 10.46 CV/g-FCW) in the four cell lines. During the long-term subcultures, the numbers of experiments used to investigate the anthocyanin biosynthetic instability within subcultures were 5, 3, 2 and 2 for cell lines A, B, C and D, respectively.
성능/효과
2. Instability of the anthocyanin accumulation of Vitis vinifera in 12 replicate flasks, originated from the same parental culture, for the lowest subculture batch variability of VC=4% (B-64) and the highest subculture variability of VC=17% CD- 63). The experiments were carried out in 250-mL flasks, containing 50 mL B5 medium, with a 5.
7% in 12 subculture batches across the four cell lines (Table 2). The highest VC variability of 17% was observed for the subculture batch, D-63, and the lowest of 4% for subculture batches, A-54, B-64 and B-65. Fig.
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