scholarly journals CRISPR-Cas9 Mediated Knockout of NtAn1 to Enhance the Lipid Accumulation in Tobacco Seed for Biodiesel Production

2020 ◽  
Author(s):  
Yinshuai Tian ◽  
Xinanbei Liu ◽  
Tingting Li ◽  
Huan Qin ◽  
Xiao Li ◽  
...  
Keyword(s):  
Protein Content ◽  
Lipid Accumulation ◽  
De Novo ◽  
Targeted Mutagenesis ◽  
Biodiesel Production ◽  
Seed Lipid ◽  
Tobacco Seed ◽  
Developing Seed ◽  
High Lipid ◽  
Increasing Demand

Abstract Background: Tobacco seed lipid is a promising non-edible feedstock for biodiesel production. In order to meet the increasing demand, achieving high seed lipid content is one of the major goals in tobacco seed production. The TT8 gene and its homologs negatively regulate seed lipid accumulation in Arabidopsis and Brassica species. We speculated that manipulating the homolog genes of TT8 in tobacco could enhance the accumulation of seed lipid.Results: In this present study, we found that the TT8 homolog genes in tobacco, NtAn1a and NtAn1b, were highly expressed in developing seed. Targeted mutagenesis of NtAn1 genes were created by the CRISPR-Cas9 based gene editing technology. Due to the defect of PAs biosynthesis, mutant seeds showed a phenotype of yellow seed coat. Seed lipid accumulation was enhanced by about 18% and 15% in two targeted mutant lines, respectively. Protein content was also significantly increased in mutant seeds. In addition, the seed yield related traits were not affected by the targeted mutagenesis of NtAn1 genes. Thus, the overall lipid productivity of the NtAn1 knockout mutants were dramatically enhanced. Conclusion: Tobacco NtAn1 genes regulate both PAs and lipid accumulation in the process of seed development. Targeted mutagenesis of NtAn1 genes could generate a yellow-seeded tobacco variety with high lipid and protein content. Furthermore, the present results revealed that CRISPR-Cas9 system could be employed in tobacco seed de novo domestication for biodiesel feedstock production.

scholarly journals Enhancement of Tobacco (Nicotiana tabacum L.) Seed Lipid Content for Biodiesel Production by CRISPR-Cas9-Mediated Knockout of NtAn1

2021 ◽  
Vol 11 ◽  
Author(s):  
Yinshuai Tian ◽  
Xinanbei Liu ◽  
Caixin Fan ◽  
Tingting Li ◽  
Huan Qin ◽  
...  
Keyword(s):  
Nicotiana Tabacum ◽  
Protein Content ◽  
Lipid Content ◽  
Lipid Accumulation ◽  
De Novo ◽  
Targeted Mutagenesis ◽  
Biodiesel Production ◽  
Seed Lipid ◽  
Tobacco Seed ◽  
Developing Seed

Tobacco (Nicotiana tabacum L.) seed lipid is a promising non-edible feedstock for biodiesel production. In order to meet the increasing demand, achieving high seed lipid content is one of the major goals in tobacco seed production. The TT8 gene and its homologs negatively regulate seed lipid accumulation in Arabidopsis and Brassica species. We speculated that manipulating the homolog genes of TT8 in tobacco could enhance the accumulation of seed lipid. In this present study, we found that the TT8 homolog genes in tobacco, NtAn1a and NtAn1b, were highly expressed in developing seed. Targeted mutagenesis of NtAn1 genes was created by the CRISPR-Cas9-based gene editing technology. Due to the defect of proanthocyanidin (PA) biosynthesis, mutant seeds showed the phenotype of a yellow seed coat. Seed lipid accumulation was enhanced by about 18 and 15% in two targeted mutant lines. Protein content was also significantly increased in mutant seeds. In addition, the seed yield-related traits were not affected by the targeted mutagenesis of NtAn1 genes. Thus, the overall lipid productivity of the NtAn1 knockout mutants was dramatically enhanced. The results in this present paper indicated that tobacco NtAn1 genes regulate both PAs and lipid accumulation in the process of seed development and that targeted mutagenesis of NtAn1 genes could generate a yellow-seeded tobacco variety with high lipid and protein content. Furthermore, the present results revealed that the CRISPR-Cas9 system could be employed in tobacco seed de novo domestication for biodiesel feedstock production.

scholarly journals Strategies for increasing lipid accumulation and recovery from Y. lipolytica: A review

OCL ◽  
2021 ◽  
Vol 28 ◽  
pp. 51
Author(s):  
Sally El Kantar ◽  
Anissa Khelfa ◽  
Eugène Vorobiev ◽  
Mohamed Koubaa
Keyword(s):  
Lipid Accumulation ◽  
Fossil Fuel ◽  
Low Cost ◽  
Operating Cost ◽  
Cost Effective ◽  
Biodiesel Production ◽  
Cultivation Conditions ◽  
Oleaginous Yeasts ◽  
Fermentation Conditions ◽  
High Lipid

Microbial-based biodiesel is produced by transesterification of lipids extracted from microbial cells, and is considered as a potential replacement of fossil fuel due to its advantages in reducing greenhouse gas emissions. Yarrowia lipolytica is one of the most studied oleaginous yeasts able to produce lipids under some fermentation conditions and is considered as a potential industrial host for biodiesel production. Several approaches have been evaluated to increase the economical attraction of biodiesel production from Y. lipolytica lipids. In this review, we highlighted the different strategies reported in the literature, allowing this yeast to achieve high lipid accumulation. These include metabolic engineering strategies, the use of low-cost effective substrates, and the optimization of the cultivation conditions for higher lipid productivity and less operating cost. We also summarized the most effective cell disruption technologies that improve the extraction efficiencies of lipids from Y. lipolytica.

Isolation and Characterization of Novel Chlorella Species with Cold Resistance and High Lipid Accumulation for Biodiesel Production

2019 ◽  
Vol 29 (6) ◽  
pp. 952-961
Author(s):  
Hyun Gi Koh ◽  
Nam Kyu Kang ◽  
Eun Kyung Kim ◽  
William Insang Suh ◽  
Won-Kun Park ◽  
...  
Keyword(s):  
Lipid Accumulation ◽  
Cold Resistance ◽  
Biodiesel Production ◽  
Isolation And Characterization ◽  
High Lipid ◽  
Chlorella Species

scholarly journals Optimization of cultivation conditions for Microcystis aeruginosa for biodiesel production using response surface methodology

2020 ◽  
Vol 44 (1) ◽  
Author(s):  
Samar A. El-Mekkawi ◽  
N. N. El-Ibiari ◽  
Ola A. El-Ardy ◽  
Nabil M. Abdelmonem ◽  
Ahmed H. Elahwany ◽  
...  
Keyword(s):  
Response Surface ◽  
Microcystis Aeruginosa ◽  
Biomass Yield ◽  
Bubble Column ◽  
Biodiesel Production ◽  
Cultivation Conditions ◽  
Inoculum Concentration ◽  
Initial Inoculum ◽  
High Lipid ◽  
Oil Concentration

Abstract Background Biodiesel is expected to play a key role in the development of a sustainable, economical, and environmentally safe source of energy. The third generation of biodiesel is derived from microalgae and cyanobacteria that have sufficient amount of oil. The optimization of biomass and oil content in biodiesel production based on algal cultivation relies upon several factors. The present experimental work aims at optimizing some of the cultivation conditions to obtain maximum oil and biomass yield and create a prediction model that describe the effect of the initial inoculum concentration, and irradiance on the biomass yield and oil concentration were designed using Design Expert 6.0.8. Results The results revealed that the optimum surface-to-volume ratio for the airlift bubble column photobioreactor was 0.9, and the most applicable model for describing Microcystis aeruginosa growth was the hyperbolic tangent model with a model constant value of 1.294 mg·L− 1·d− 1/μmol·m− 2·s− 1. The optimum cultivation conditions were 81 μmol·m− 2·s− 1 irradiance and 67 mg·L− 1 initial inoculum concentration, and these conditions achieved a biomass yield of 163 mg·L− 1·d− 1 and an oil concentration of 143 mg·L− 1. Conclusions This work focused on the cultivation of microalgae in closed systems. Cyanobacteria as M. aeruginosa has high lipid content, and high lipid productivity makes it suitable as a lipid feed stock for biodiesel production. The response surface method was the most suitable route to study the simultaneous influence of irradiance and initial inoculum concentration through statistical methods as well as to establish a model for predicting the biomass yield and oil concentration of M. aeruginosa.

scholarly journals Snf1 Is a Regulator of Lipid Accumulation in Yarrowia lipolytica

2013 ◽  
Vol 79 (23) ◽  
pp. 7360-7370 ◽  
Author(s):  
John Seip ◽  
Raymond Jackson ◽  
Hongxian He ◽  
Quinn Zhu ◽  
Seung-Pyo Hong
Keyword(s):  
Yarrowia Lipolytica ◽  
Lipid Accumulation ◽  
Oleaginous Yeast ◽  
De Novo ◽  
Lipid Synthesis ◽  
Omega 3 ◽  
Wild Type ◽  
Content Type ◽  
Reverse Transcription Pcr ◽  
Dry Cell Weight

ABSTRACTIn the oleaginous yeastYarrowia lipolytica,de novolipid synthesis and accumulation are induced under conditions of nitrogen limitation (or a high carbon-to-nitrogen ratio). The regulatory pathway responsible for this induction has not been identified. Here we report that the SNF1 pathway plays a key role in the transition from the growth phase to the oleaginous phase inY. lipolytica. Strains with aY. lipolyticasnf1(Ylsnf1) deletion accumulated fatty acids constitutively at levels up to 2.6-fold higher than those of the wild type. When introduced into aY. lipolyticastrain engineered to produce omega-3 eicosapentaenoic acid (EPA),Ylsnf1deletion led to a 52% increase in EPA titers (7.6% of dry cell weight) over the control. Other components of theY. lipolyticaSNF1 pathway were also identified, and their function in limiting fatty acid accumulation is suggested by gene deletion analyses. Deletion of the gene encoding YlSnf4, YlGal83, or YlSak1 significantly increased lipid accumulation in both growth and oleaginous phases compared to the wild type. Furthermore, microarray and quantitative reverse transcription-PCR (qRT-PCR) analyses of theYlsnf1mutant identified significantly differentially expressed genes duringde novolipid synthesis and accumulation inY. lipolytica. Gene ontology analysis found that these genes were highly enriched with genes involved in lipid metabolism. This work presents a new role for Snf1/AMP-activated protein kinase (AMPK) pathways in lipid accumulation in this oleaginous yeast.

scholarly journals 905 FASN prevents immunogenicity of irradiated glioblastoma by inhibiting ER stress

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A950-A950
Author(s):  
Mara De Martino ◽  
Camille Daviaud ◽  
Claire Vanpouille-Box
Keyword(s):  
Er Stress ◽  
Lipid Accumulation ◽  
Fatty Acid Synthase ◽  
Immune Escape ◽  
De Novo ◽  
Lipid Synthesis ◽  
Adult Brain ◽  
Immune Recognition

BackgroundGlioblastoma (GBM) is the most aggressive and incurable adult brain tumor. Radiation therapy (RT) is an essential modality for GBM treatment and is recognized to stimulate anti-tumor immunity by inducing immunogenic cell death (ICD) subsequent to endoplasmic reticulum (ER) stress. However, RT also exacerbates potent immunosuppressive mechanisms that facilitate immune evasion. Notably, increased de novo lipid synthesis by the fatty acid synthase (FASN) is emerging as a mechanism of therapy resistance and immune escape. Here, we hypothesize that RT induces FASN to promote GBM survival and evade immune recognition by inhibiting ER stress and ICD.MethodsTo determine if lipid synthesis is altered in response to RT, we first assessed FASN expression by western blot (WB) and lipid accumulation by BODIPY staining in murine (CT2A and GL261) and human (U118) GBM cell lines. Next, FASN expression was blocked in CT2A cells using CRISPR-Cas9 or an inducible shRNA directed against Fasn to evaluate ICD and ER stress markers by ELISA, WB, and electron microscopy. Finally, CT2AshFASN cells or its non-silencing control (CT2AshNS) were orthotopically implanted and FASN knockdown was induced by feeding the mice with doxycycline. The immune contexture was determined by in situ immunofluorescence (n=3/group). Remaining mice were followed for survival (n=7/group).ResultsWe found that in vitro irradiation of GBM cells induces lipid accumulation in a dose-dependent fashion; an effect that is magnified over time lasting at least 6/7 days. Consistent with these findings, FASN expression was upregulated in irradiated GBM cells. Confirming the role of FASN, RT-induced accumulation of lipids was reverted when GBM cells were incubated with a FASN inhibitor. Next, we found that FASN ablation in CT2A cells induces mitochondria disruption and was sufficient to increase the expression of the ER stress makers BIP and CHOP. Along similar lines, shFASN enhances the secretion of the ICD markers HMGB1, IFN-beta and CXCL10 in irradiated CT2A cells. In vivo, CT2AshFASN tumors presented increased infiltration of CD11c+ cells and CD8+ T cells, consistent with prolonged mice survival (56 days vs. 28 days for CT2AshNS). Importantly, 43% of CT2AshFASN-bearing mice remained tumor-free for more than 70 days, while none of the CT2AshNS-bearing mice survived.ConclusionsAltogether, our data suggest that FASN-mediated lipid synthesis is an important mechanism to prevent ER stress, ICD, and anti-tumor immune responses in GBM. While much work remains to be done, our data propose FASN as a novel therapeutic target to overcome immunosuppression and sensitize GBM to immunotherapies.

scholarly journals Initial study of lipid accumulation in green algal Haematococcus pluvialis Flotow cultured in liquid Bold’s Basal medium aerated

2019 ◽  
Vol 3 (3) ◽  
pp. 144-149
Author(s):  
Nguyen Tran Dong Phuong ◽  
Le Huyen Ai Thuy ◽  
Bui Trang Viet
Keyword(s):  
Lipid Accumulation ◽  
Fatty Acid Biosynthesis ◽  
Haematococcus Pluvialis ◽  
Acyl Carrier Protein ◽  
Nile Red ◽  
Basal Medium ◽  
Initial Study ◽  
Rt Pcr ◽  
Green Algal ◽  
High Lipid

The fresh green algal Haematococcus pluvialis Flotow was proved to be the starting material for the production of biofuel, high lipid content along with astaxanthin, a high value colorant. In this study, lipid accumulation in H. pluvialis cultured in liquid Bold’s Basal medium aerated was investigated for a period of 12 weeks. Lipid accumulation was evaluated through the expression of two genes: BC (biotin carboxylase, initial gene) and FATA (acyl-acyl carrier protein thioesterase, end gene) in the process of fatty acid biosynthesis with Real-time RT-PCR, lipid determination by Nile Red and biodiesel quantifying by transesterification. The results showed that the expression of two BC and FATA genes was recorded at all weeks of culture. However, the expression of BC and FATA genes increased gradually from the week 9 (1.3, 4.1, respectively) to week 11 (1.7, 30.9, respectively). Meanwhile, yellow fluorescence in the microalgal cells showed that lipid appeared from week 6 to week 12. The obtained biodiesel increased slowly from week 8 (0.036 mg/mL) to week 12 (0.041 mg/mL). At week 11, the expression values of both BC gene (1.7) and FATA gene (30.9) were maximized, leading to the highest biodiesel content at the week 12.

scholarly journals Effects of nitrogen, phosphorus, EDTA and sodium chloride on biomass and lipid accumulation of Chaetomorpha aerea

2020 ◽  
pp. 152-158
Author(s):  
Gour Gopal Satpati ◽  
Ruma Pal
Keyword(s):  
Fatty Acids ◽  
Sodium Chloride ◽  
Total Lipid ◽  
Lipid Accumulation ◽  
Saturated Fatty Acids ◽  
Stress Conditions ◽  
Biodiesel Production ◽  
Cultural Conditions ◽  
Chaetomorpha Aerea ◽  
Nitrogen Phosphorus

The increase of total lipid and fatty acids production was studied under different nutrient stress conditions using the macroalga, Chaetomorpha aerea. The effects of nitrogen, phosphorus, ethylene diamine tetra-acetate and sodium chloride on the growth and lipid accumulation were systematically investigated in laboratory conditions. The biomass was harvested at different stages of cultivation and assessed. The maximum changes of growth and lipid accumulation were observed in the exponential phase at different cultural conditions. The two-fold increase of total lipid was found in the order of 28.27±0.04 % (at 0.1 g/L nitrogen) > 27.30±0.37 % (at 0.5 g/L of phosphorus) > 25.86±0.77 % (at 0.05 g/L of EDTA)> 24.37±0.04 % (at 0.05 g/L NaCl) on 8th day of cultivation. The fatty acids were identified and quantified by gas chromatography mass spectrometry (GC-MS). The alga produces significantly high amount of monounsaturated fatty acid (MUFA) and saturated fatty acids (SFA) than the polyunsaturated fatty acids (PUFA) in different cultural conditions. The elevated levels of C16:1, C18:1 and C20:1 was identified under nitrate, phosphate and salt stress conditions, which are more suitable for biodiesel production.

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