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Xiangming Li Yujian Zhang Li Jing Zongming Fu Ou Ma Jishna Ganguly Nilesh Vaidya Richard Sisson Jennifer Naginskaya Avinash Chinthala Minggang Cui Ryan Yamagata Mark Wilson Matthew Sanders Zihao Wang Paola Lo Surdo Marcin Bugno 《Biotechnology progress》2020,36(2):e2914
Mammalian cell line generation typically includes stable pool generation, single cell cloning and several rounds of clone selection based on cell growth, productivity and product quality criteria. Individual clone expansion and phenotype-based ranking is performed initially for hundreds or thousands of mini-scale cultures, representing the major operational challenge during cell line development. Automated cell culture and analytics systems have been developed to enable high complexity clone selection workflows; while ensuring traceability, safety, and quality of cell lines intended for biopharmaceutical applications. Here we show that comprehensive and quantitative assessment of cell growth, productivity, and product quality attributes are feasible at the 200–1,200 cell colony stage, within 14 days of the single cell cloning in static 96-well plate culture. The early cell line characterization performed prior to the clone expansion in suspension culture can be used for a single-step, direct selection of high quality clones. Such clones were comparable, both in terms of productivity and critical quality attributes (CQAs), to the top-ranked clones identified using an established iterative clone screening approach. Using a complex, multi-subunit antigen as a model protein, we observed stable CQA profiles independently of the cell culture format during the clonal expansion as well as in the batch and fed-batch processes. In conclusion, we propose an accelerated clone selection approach that can be readily incorporated into various cell line development workstreams, leading to significant reduction of the project timelines and resource requirements. 相似文献
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Leaf segments excised from Centella asiatica, a medicinal and neutraceutical plant, produced abundant somaticembryoswhen cultured onMS mediumwith 9.29 Mkinetin in combination with 2.26 M2,4-D. Granular, white,shiny clusters of callus developed after 1 week of culture, and then formed heart and cotyledonary stage embryoson the same medium after 4 weeks. Somatic embryos matured and germinated in the presence of MS mediumcontaining 2.32 M kinetin with (2.89M) GA3. Plantlets were successfully transferred to pots containing amixture of soil and vermiculite (1:1). 相似文献
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Plant peroxidases are one of the most extensively studied group of enzymes which find applications in the environment, health,
pharmaceutical, chemical and biotechnological processes. Class III secretary peroxidase from alfalfa (Medicago sativa) has been
characterized using bioinformatics approach Physiochemical properties and topology of alfalfa peroxidase were compared with
that of soybean and horseradish peroxidase, two most popular commercially available peroxidase preparations. Lower value of
instability index as predicted by ProtParam and presence of extra disulphide linkages as predicted by Cys_REC suggested alfalfa
peroxidase to be more stable than either of the commercial preparations. Multiple Sequence Alignment (MSA) with other
functionally similar proteins revealed the presence of highly conserved catalytic residues. Three dimensional model of alfalfa
peroxidase was constructed based on the crystal structure of soybean peroxidase (PDB Id: 1FHF A) by homology modelling
approach. The model was checked for stereo chemical quality by PROCHECH, VERIFY 3D, WHAT IF, ERRAT, 3D MATCH AND
ProSA servers. The best model was selected, energy minimized and used to analyze structure function relationship with substrate
hydrogen peroxide by Autodock 4.0. The enzyme substrate complex was viewed with Swiss PDB viewer and one residue ASP43
was found to stabilize the interaction by hydrogen bonds. The results of the study may be a guiding point for further investigations
on alfalfa peroxidase. 相似文献
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