Electroporation Procedures for Genetic Modification of Green Algae (Chlorella spp.)

Ji, Liang; Fan, Jianhua

doi:10.1007/978-1-4939-9740-4_20

Cited by 10 publications

(6 citation statements)

References 10 publications

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“…reinhardtii , [ 67–73 ] D . salina , [ 74 ] Chlorella spp., [ 75–79 ] Scenedesmus obliquus , [ 80 ] Neochloris oleoabundans , [ 81 ] and Nannochloropsis spp. [ 82–85 ]…”

Section: Mutagenesis and Genetic Bioengineering Of Microalgaementioning

confidence: 99%

“…Electroporation is a physical gene transfer method that uses highvoltage pulses to generate temporary pores on the membrane to enter genes into intracellular space of protoplasts, wall-less cells, or even the cells with rigid cell walls microalgae. Successful transformation by electroporation has been operated in numerous microalgae like C. reinhardtii, [67][68][69][70][71][72][73] D. salina, [74] Chlorella spp., [75][76][77][78][79] Scenedesmus obliquus, [80] Neochloris oleoabundans, [81] and Nannochloropsis spp. [82][83][84][85]…”

Section: Electroporationmentioning

confidence: 99%

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Multiomics approaches and genetic engineering of metabolism for improved biorefinery and wastewater treatment in microalgae

Kuo

Yang

Chien

et al. 2022

Biotechnology Journal

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Microalgae, a group of photosynthetic microorganisms rich in diverse and novel bioactive metabolites, have been explored for the production of biofuels, high value-added compounds as food and feeds, and pharmaceutical chemicals as agents with therapeutic benefits. This article reviews the development of omics resources and genetic engineering techniques including gene transformation methodologies, mutagenesis, and genome-editing tools in microalgae biorefinery and wastewater treatment (WWT).The introduction of these enlisted techniques has simplified the understanding of complex metabolic pathways undergoing microalgal cells. The multiomics approach of the integrated omics datasets, big data analysis, and machine learning for the discovery of objective traits and genes responsible for metabolic pathways was reviewed. Recent advances and limitations of multiomics analysis and genetic bioengineering technology to facilitate the improvement of microalgae as the dual role of WWT and biorefinery feedstock production are discussed.

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“…reinhardtii , [ 67–73 ] D . salina , [ 74 ] Chlorella spp., [ 75–79 ] Scenedesmus obliquus , [ 80 ] Neochloris oleoabundans , [ 81 ] and Nannochloropsis spp. [ 82–85 ]…”

Section: Mutagenesis and Genetic Bioengineering Of Microalgaementioning

confidence: 99%

Section: Electroporationmentioning

confidence: 99%

Multiomics approaches and genetic engineering of metabolism for improved biorefinery and wastewater treatment in microalgae

Kuo

Yang

Chien

et al. 2022

Biotechnology Journal

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“…Two methods were employed for the chloroplast transformation in C. vulgaris using pulse field electroporation. The first methodology was based on the procedure reported by Ji and Fan (2020). The microalgal cells were centrifuged, and the pellet was washed in ice-cold hypertonic buffer solution containing 0.2 M mannitol and 0.2 M sorbitol for 1 h by immersion in ice.…”

Section: Chloroplast Transformation In C Vulgaris By Electroporationmentioning

confidence: 99%

Construction and Validation of a Chloroplast Expression Vector for the Production of Recombinant Proteins in Chlorella vulgaris

Bolaños-Martínez¹,

Malla²,

Rosales‐Mendoza³

et al. 2022

Front. Mar. Sci.

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Microalgae constitute a diverse group of photosynthetic unicellular microorganisms that have gained immense attention for biotechnological applications. They are promising platforms for the production of high-value metabolites and biopharmaceuticals for commercial and therapeutic applications because of their physiological properties and capability to grow easily in natural and artificial environments. Although the proof-of-concept for some applications have been achieved for model species, such as Chlamydomonas reinhardtii, the genetic engineering methods for microalgae are still in their infancy. Thus, an expansion of this field is required. Chlorella vulgaris is an important algal species with a high protein content and requires focus for the development of an efficient nuclear and chloroplast transformation process. This research aimed to establish a chloroplast transformation method for the freshwater green-algae species C. vulgaris based on a specific expression vector (pCMCC, which was named after Chula Mexico Chlorella chloroplast) constructed with endogenous recombination regions, namely, 16S–trn I (left) and trn A–23S (right), and the Prrn promoter. Human basic fibroblast growth factor (bFGF) was adopted as a target biopharmaceutical to establish the chloroplast expression approach. The plasmid pCMCC:bFGF was transformed into C. vulgaris via electroporation using simple carbohydrate-based buffers, which aided in the transfer of the transgene into the chloroplast genome. Cells transformed with the pCMCC:bFGF vector were selected using kanamycin, and resistant colonies were analyzed by polymerase chain reaction and Western blotting to confirm the presence of the transgene and the recombinant bFGF, respectively. The bFGF that accumulated in the transplastomic C. vulgaris clones ranged from 0.26 to 1.42 ng/g fresh weight of biomass, and it was quantified by enzyme-linked immunosorbent assay. Therefore, the designed expression vector, in combination with an optimized electroporation protocol, constitutes a viable approach to successfully develop transplastomic lines of C. vulgaris for the potential low-cost production of biopharmaceuticals using this algal species. This study paves the way for the establishment of chloroplast biotechnology in microalgae other than the model organism C. reinhardtii.

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“…108,109 Recently, microalgal transformation techniques through microelectroporation have evolved to improve the delivery of exogenous molecules in microalgae. [110][111][112] Digital microdroplet electroporation has been developed for gene transformation of microalgae in a microuidic system (Fig. 6a).…”

Section: Electroporation For Microalgal Transformationmentioning

confidence: 99%

“…108,109 Recently, microalgal transformation techniques through micro-electroporation have evolved to improve the delivery of exogenous molecules in microalgae. 110–112…”

Section: Recent Advances In Electroporation For Microalgal Biodiesel ...mentioning

confidence: 99%

Recent advances in photobioreactor systems for sustainable and enhanced microalgal biofuel production

Kim

Yoo

Lee

2022

Sustainable Energy Fuels

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Electroporation Procedures for Genetic Modification of Green Algae (Chlorella spp.)

Cited by 10 publications

References 10 publications

Multiomics approaches and genetic engineering of metabolism for improved biorefinery and wastewater treatment in microalgae

Multiomics approaches and genetic engineering of metabolism for improved biorefinery and wastewater treatment in microalgae

Construction and Validation of a Chloroplast Expression Vector for the Production of Recombinant Proteins in Chlorella vulgaris

Recent advances in photobioreactor systems for sustainable and enhanced microalgal biofuel production

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