Improved expression and purification of recombinant human serum albumin from transgenic tobacco suspension culture

Sun, Qiao-Yang; Ding, Ling-Wen; Lomonossoff, George P.; Sun, Yongbing; Luo, Ming; Li, Chaoqiong; Jiang, Liwen; Xu, Zeng-Fu

doi:10.1016/j.jbiotec.2011.06.033

Cited by 48 publications

(26 citation statements)

References 43 publications

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“…Tobacco Bright Yellow-2 cells stably transformed with pEAQ-SP- HSA expressed recombinant human serum albumin (rHSA) at 11.88 mg/L in the culture medium, accounting for 0.7% TSP. In contrast a conventional CaMV 35S-based vector (lacking P19) failed to yield detectable quantities of secreted rHSA [44]. …”

Section: Resultsmentioning

confidence: 99%

RNA viral vectors for improved Agrobacterium-mediated transient expression of heterologous proteins in Nicotiana benthamiana cell suspensions and hairy roots

Larsen

Curtis

2012

BMC Biotechnol

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BackgroundPlant cell suspensions and hairy root cultures represent scalable protein expression platforms. Low protein product titers have thus far limited the application of transient protein expression in these hosts. The objective of this work was to overcome this limitation by harnessing A. tumefaciens to deliver replicating and non-replicating RNA viral vectors in plant tissue co-cultures.ResultsReplicating vectors derived from Potato virus X (PVX) and Tobacco rattle virus (TRV) were modified to contain the reporter gene β-glucuronidase (GUS) with a plant intron to prevent bacterial expression. In cell suspensions, a minimal PVX vector retaining only the viral RNA polymerase gene yielded 6.6-fold more GUS than an analogous full-length PVX vector. Transient co-expression of the minimal PVX vector with P19 of Tomato bushy stunt virus or HC-Pro of Tobacco etch virus to suppress post-transcriptional gene silencing increased GUS expression by 44 and 83%, respectively. A non-replicating vector containing a leader sequence from Cowpea mosaic virus (CPMV-HT) modified for enhanced translation led to 70% higher transient GUS expression than a control treatment. In hairy roots, a TRV vector capable of systemic movement increased GUS accumulation by 150-fold relative to the analogous PVX vector. Histochemical staining for GUS in TRV-infected hairy roots revealed the capacity for achieving even higher productivity per unit biomass.ConclusionsFor the first time, replicating PVX vectors and a non-replicating CPMV-HT vector were successfully applied toward transient heterologous protein expression in cell suspensions. A replicating TRV vector achieved transient GUS expression levels in hairy roots more than an order of magnitude higher than the highest level previously reported with a viral vector delivered by A. tumefaciens.

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Section: Resultsmentioning

confidence: 99%

RNA viral vectors for improved Agrobacterium-mediated transient expression of heterologous proteins in Nicotiana benthamiana cell suspensions and hairy roots

Larsen

Curtis

2012

BMC Biotechnol

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“…By contrast, RNA virus-based vector systems, such as those using TMV or Potato virus X incorporate both the relevant viral replicase and movement proteins, and this results in severe host range limitations, essentially confining these systems to the Solanaceae. Similarly, high expression afforded by the nonreplicating, hypertranslation system described by Sainsbury and Lomonossoff (2008), has been limited to members of the Nicotiana genus (Sainsbury and Lomonossoff, 2008;Sun et al, 2011).…”

Section: Discussionmentioning

confidence: 99%

In Plant Activation: An Inducible, Hyperexpression Platform for Recombinant Protein Production in Plants

et al. 2013

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“…Strategies for plant transformation contain stable nuclear transformation, stable plastid transformation, plant cell-suspension, and transient expression systems [5]. Plant cell suspension cultures have several advantages including the capacity for shorter life cycles, independence from environmental effects such as climate, soil quality, season, day length and weather, the lack of biosafety issues such as gene flow via pollen, and the possibility of bacterial contamination from the plant growth environment [6]. But, the yield and quality of recombinant proteins in plant cell culture-based expression systems need to be further improved.…”

Section: Introductionmentioning

confidence: 99%

Stable Plastid Transformation for High-Level Recombinant Protein Expression: Promises and Challenges

Gao

Xue

et al. 2012

Journal of Biomedicine and Biotechnology

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Plants are a promising expression system for the production of recombinant proteins. However, low protein productivity remains a major obstacle that limits extensive commercialization of whole plant and plant cell bioproduction platform. Plastid genetic engineering offers several advantages, including high levels of transgenic expression, transgenic containment via maternal inheritance, and multigene expression in a single transformation event. In recent years, the development of optimized expression strategies has given a huge boost to the exploitation of plastids in molecular farming. The driving forces behind the high expression level of plastid bioreactors include codon optimization, promoters and UTRs, genotypic modifications, endogenous enhancer and regulatory elements, posttranslational modification, and proteolysis. Exciting progress of the high expression level has been made with the plastid-based production of two particularly important classes of pharmaceuticals: vaccine antigens, therapeutic proteins, and antibiotics and enzymes. Approaches to overcome and solve the associated challenges of this culture system that include low transformation frequencies, the formation of inclusion bodies, and purification of recombinant proteins will also be discussed.

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Improved expression and purification of recombinant human serum albumin from transgenic tobacco suspension culture

Cited by 48 publications

References 43 publications

RNA viral vectors for improved Agrobacterium-mediated transient expression of heterologous proteins in Nicotiana benthamiana cell suspensions and hairy roots

RNA viral vectors for improved Agrobacterium-mediated transient expression of heterologous proteins in Nicotiana benthamiana cell suspensions and hairy roots

In Plant Activation: An Inducible, Hyperexpression Platform for Recombinant Protein Production in Plants

Stable Plastid Transformation for High-Level Recombinant Protein Expression: Promises and Challenges

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