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Enhancement of glycerol metabolism in the oleaginous marine diatom Fistulifera solaris JPCC DA0580 to improve triacylglycerol productivity.

Muto M, Tanaka M, Liang Y, Yoshino T, Matsumoto M, Tanaka T - Biotechnol Biofuels (2015)

Bottom Line: Overexpression of the endogenous glycerol kinase (GK) gene in an oleaginous marine diatom, Fistulifera solaris JPCC DA0580, accelerates glycerol metabolism and improves lipid and biomass productivities.We have demonstrated the potential of metabolic engineering in oleaginous microalgae to improve lipid productivity.Metabolic engineering techniques can be used to optimize BDF production.

View Article: PubMed Central - PubMed

Affiliation: Division of Biotechnology and Life Science, Institute of Engineering, Tokyo University of Agriculture and Technology, 2-24-16 Naka-cho, Koganei, Tokyo 184-8588 Japan ; JST, CREST, Sanbancho 5, Chiyoda-ku, Tokyo 102-0075 Japan.

ABSTRACT

Background: Microalgal oil is a promising alternative feedstock for biodiesel fuel (BDF). Mixotrophic cultivation with glycerol, the primary byproduct of BDF production, may be used to optimize BDF production. This strategy would reduce costs through glycerol recycling and improve lipid productivity and biomass productivity by overcoming the growth retardation caused by decreased light penetration in high-density culture.

Results: Overexpression of the endogenous glycerol kinase (GK) gene in an oleaginous marine diatom, Fistulifera solaris JPCC DA0580, accelerates glycerol metabolism and improves lipid and biomass productivities. Two candidates were selected from a collection of 90 G418-resistant clones, based on growth and confirmation of genome integration. GK gene expression was higher in the selected clones (GK1_7 and GK2_16) than in the wild-type culture. The GK2_16 clone achieved a 12% increase in lipid productivity.

Conclusion: We have demonstrated the potential of metabolic engineering in oleaginous microalgae to improve lipid productivity. Metabolic engineering techniques can be used to optimize BDF production.

No MeSH data available.


Related in: MedlinePlus

Heterologous gene detection inFistulifera solaristransformed with pSP-GK1/fcpB or pSP-GK2/fcpB. The nptII(A) and nptII-glycerol kinase (GK) gene fragments (B) were PCR-amplified. M, 1-kb marker; P, pSP-GK1/fcpB as the positive control; W, wild type; N, negative control transformed with pSP-NPT/H4; GK1, transformed with pSP-GK1/fcpB; GK2, transformed with pSP-GK2/fcpB.
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Fig2: Heterologous gene detection inFistulifera solaristransformed with pSP-GK1/fcpB or pSP-GK2/fcpB. The nptII(A) and nptII-glycerol kinase (GK) gene fragments (B) were PCR-amplified. M, 1-kb marker; P, pSP-GK1/fcpB as the positive control; W, wild type; N, negative control transformed with pSP-NPT/H4; GK1, transformed with pSP-GK1/fcpB; GK2, transformed with pSP-GK2/fcpB.

Mentions: The presence of the integrated genes in the 10 selected clones was verified by PCR (FigureĀ 2). A 795-bp fragment encoding the nptII gene region was amplified from the genomic DNA of G418-resistant clones. All 10 clones, as well as the negative control with the empty pSP-NPT/H4 vector, showed the presence of the nptII gene, while no amplified product was obtained from the wild type. On the other hand, the combined fragment of the nptII gene and the GK1 or GK2 gene region (2622 bp) was amplified from only three of the clones (GK1_7, GK2_16, and GK2_39). The wild type and the negative control showed no nptII-GK product. These results suggest that partial gene integration of the vector may occur during transformation in F. solaris. The GK2_39 clone showed a decrease in growth rate (Additional file 1) and was eliminated from subsequent experiments.Figure 2


Enhancement of glycerol metabolism in the oleaginous marine diatom Fistulifera solaris JPCC DA0580 to improve triacylglycerol productivity.

Muto M, Tanaka M, Liang Y, Yoshino T, Matsumoto M, Tanaka T - Biotechnol Biofuels (2015)

Heterologous gene detection inFistulifera solaristransformed with pSP-GK1/fcpB or pSP-GK2/fcpB. The nptII(A) and nptII-glycerol kinase (GK) gene fragments (B) were PCR-amplified. M, 1-kb marker; P, pSP-GK1/fcpB as the positive control; W, wild type; N, negative control transformed with pSP-NPT/H4; GK1, transformed with pSP-GK1/fcpB; GK2, transformed with pSP-GK2/fcpB.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4308894&req=5

Fig2: Heterologous gene detection inFistulifera solaristransformed with pSP-GK1/fcpB or pSP-GK2/fcpB. The nptII(A) and nptII-glycerol kinase (GK) gene fragments (B) were PCR-amplified. M, 1-kb marker; P, pSP-GK1/fcpB as the positive control; W, wild type; N, negative control transformed with pSP-NPT/H4; GK1, transformed with pSP-GK1/fcpB; GK2, transformed with pSP-GK2/fcpB.
Mentions: The presence of the integrated genes in the 10 selected clones was verified by PCR (FigureĀ 2). A 795-bp fragment encoding the nptII gene region was amplified from the genomic DNA of G418-resistant clones. All 10 clones, as well as the negative control with the empty pSP-NPT/H4 vector, showed the presence of the nptII gene, while no amplified product was obtained from the wild type. On the other hand, the combined fragment of the nptII gene and the GK1 or GK2 gene region (2622 bp) was amplified from only three of the clones (GK1_7, GK2_16, and GK2_39). The wild type and the negative control showed no nptII-GK product. These results suggest that partial gene integration of the vector may occur during transformation in F. solaris. The GK2_39 clone showed a decrease in growth rate (Additional file 1) and was eliminated from subsequent experiments.Figure 2

Bottom Line: Overexpression of the endogenous glycerol kinase (GK) gene in an oleaginous marine diatom, Fistulifera solaris JPCC DA0580, accelerates glycerol metabolism and improves lipid and biomass productivities.We have demonstrated the potential of metabolic engineering in oleaginous microalgae to improve lipid productivity.Metabolic engineering techniques can be used to optimize BDF production.

View Article: PubMed Central - PubMed

Affiliation: Division of Biotechnology and Life Science, Institute of Engineering, Tokyo University of Agriculture and Technology, 2-24-16 Naka-cho, Koganei, Tokyo 184-8588 Japan ; JST, CREST, Sanbancho 5, Chiyoda-ku, Tokyo 102-0075 Japan.

ABSTRACT

Background: Microalgal oil is a promising alternative feedstock for biodiesel fuel (BDF). Mixotrophic cultivation with glycerol, the primary byproduct of BDF production, may be used to optimize BDF production. This strategy would reduce costs through glycerol recycling and improve lipid productivity and biomass productivity by overcoming the growth retardation caused by decreased light penetration in high-density culture.

Results: Overexpression of the endogenous glycerol kinase (GK) gene in an oleaginous marine diatom, Fistulifera solaris JPCC DA0580, accelerates glycerol metabolism and improves lipid and biomass productivities. Two candidates were selected from a collection of 90 G418-resistant clones, based on growth and confirmation of genome integration. GK gene expression was higher in the selected clones (GK1_7 and GK2_16) than in the wild-type culture. The GK2_16 clone achieved a 12% increase in lipid productivity.

Conclusion: We have demonstrated the potential of metabolic engineering in oleaginous microalgae to improve lipid productivity. Metabolic engineering techniques can be used to optimize BDF production.

No MeSH data available.


Related in: MedlinePlus