Uso de tecnologías CRISPR-CAS9 en microalgas aplicado a la obtención de productos biotecnológicos de interés industrial

Resumen

Las microalgas se destacan por su capacidad de adaptarse a diferentes medios de cultivo, participación en captura de CO2  y productos de interés biotecnológico cómo biocombustibles, proteínas, biofertilizantes, suplementos alimenticios, pigmentos, entre otros. Se han trabajado diferentes tipos de estrés como variación del pH, deficiencia de nutrientes, estrés salino, cambios en la temperatura y alta irradiancia con el fin de aumentar la producción de metabolitos aplicado a diferentes industrias; sin embargo, la demanda de los productos biotecnológicos se ha incrementado ante las necesidades de la población, por lo que el uso de tecnologías de modificación genética ha surgido como una alternativa  durante la última década gracias a la capacidad y eficiencia que muestran los métodos como mutagénesis aleatoria, supresión de genes y crispr-cas9. Se propone como objetivo conocer la aplicación del crispr-cas9 en microalgas relacionado con la obtención de productos biotecnológicos de interés industrial. Se determinó que esta tecnología aplicada en las microalgas con uso industrial incrementa la obtención de productos de interés biotecnológico como lípidos, carotenoides, proteínas y enzimas recombinantes.

Palabras clave:

microalga CRISPR-CAS9 Metabolitos Productos biotecnologicos Tecnologías de modificación genética ciencias naturales

Citas

Ajjawi, I., Verruto, J., Aqui, M., Soriaga, L. B., Coppersmith, J., Kwok, K., Peach, L., Orchard, E., Kalb, R., Xu, W., Carlson, T. J., Francis, K., Konigsfeld, K., Bartalis, J., Schultz, A., Lambert, W., Schwartz, A. S., Brown, R., & Moellering, E. R. (2017). Lipid production in Nannochloropsis gaditana is doubled by decreasing expression of a single transcriptional regulator. Nature Biotechnology, 35(7), 647-652. https://doi.org/10.1038/nbt.3865

Ahirwar, A., Meignen, G., Khan, M. J., Sirotiya, V., Scarsini, M., Roux, S., ... Vinayak, V. (2021). Light modulates transcriptomic dynamics upregulating astaxanthin accumulation in Haematococcus: A review. Bioresource Technology, 340, 125707. https://doi.org/10.1016/j.biortech.2021.125707

Anjos, M., Fernandes, B. D., Vicente, A. A., Teixeira, J. A., & Dragone, G. (2013). Optimization of CO2 bio-mitigation by Chlorella vulgaris. Bioresource Technology, 139, 149-154. https://doi.org/10.1016/j.biortech.2013.04.032

Araujo Abad, S. y Collahuazo Reinoso, Y. (2019). Producción de Biofertilizantes a Partir de Microalgas. CEDAMAZ, 9(2), 81–87.

Baek, K., Kim, D. H., Jeong, J., Sim, S. J., Melis, A., Kim, J., Jin, E. & Bae, S. (2016). DNA-free two-gene knockout in Chlamydomonas reinhardtii via CRISPR-Cas9 ribonucleoproteins. Scientific Reports, 6(1), 30620. https://doi.org/10.1038/srep30620

Bernheim, A., Calvo-Villamañán, A., Basier, C., Cui, L., Rocha, E. P. C., Touchon, M., & Bikard, D. (2017). Inhibition of NHEJ repair by type II-A CRISPR-Cas systems in bacteria. Nature Communications, 8(1), 2094-9. https://doi.org/10.1038/s41467-017-02350-1

Bharathkumar, N., Sunil, A., Meera, P., Aksah, S., Kannan, M., Saravanan, K. M., & Anand, T. (2022). CRISPR/Cas-Based Modifications for Therapeutic Applications: A Review. Molecular Biotechnology, 64(4), 355-372. https://doi.org/10.1007/s12033-021-00422-8

Burmistrz, M., Krakowski, K., & Krawczyk-Balska, A. (2020). RNA-Targeting CRISPR–Cas Systems and Their Applications. International Journal of Molecular Sciences, 21(3), 1122. https://doi.org/10.3390/ijms21031122

Camacho Kurmen, J. E., González, G. y Klotz, B. (2013). Producción de Astaxantina en Haematococcus pluvialis bajo diferentes condiciones de estrés. Nova, 11(19), 93. https://doi.org/10.22490/24629448.1022

Castillo, O., Torres-Badajoz, S., Núñez-Colín, C., Peña-Caballero, V., Herrera Méndez, C. y Rodríguez-Núñez, J. (2017). Producción de biodiésel a partir de microalgas: avances y perspectivas biotecnológicas. Hidrobiológica, 27(3), 337-352. https://search.proquest.com/docview/2097445304

Chang, K. S., Kim, J., Park, H., Hong, S., Lee, C., & Jin, E. (2020). Enhanced lipid productivity in AGP knockout marine microalga Tetraselmis sp. using a DNA-free CRISPR-Cas9 RNP method. Bioresource Technology, 303, 122932. https://doi.org/10.1016/j.biortech.2020.122932

Chaumont, D. & Thépenier, C. (1995). Carotenoid content in growing cells of Haematococcus pluvialis during a sunlight cycle. Journal of applied phycology, 7, 529-537

Chen, E., Lin-Shiao, E., Trinidad, M., Saffari Doost, M., Colognori, D., & Doudna, J. A. (2022). Decorating chromatin for enhanced genome editing using CRISPR-Cas9. Nevada RNformation, 119(49), e2204259119. https://doi.org/10.1073/pnas.2204259119

Coronado-Reyes, J. A., Salazar-Torres, J. A., Juárez-Campos, B., & González-Hernández, J. C. (2022). Chlorella vulgaris, a microalgae important to be used in Biotechnology: a review. Ciência E Tecnología De Alimentos, 42. https://doi.org/10.1590/fst.37320

Dalvie, N. C., Lorgeree, T., Biedermann, A. M., Love, K. R., & Love, J. C. (2022). Simplified Gene Knockout by CRISPR-Cas9-Induced Homologous Recombination. ACS Synthetic Biology, 11(1), 497-501. https://doi.org/10.1021/acssynbio.1c00194

Das, D. (2016). Engineering Spirulina for Enhanced Medicinal Application. Algal Biorefinery: An Integrated Approach (pp. 235-252). Springer International Publishing AG. https://doi.org/10.1007/978-3-319-22813-6_11

Dhamad, A. E., & Lessner, D. J. (2020). A CRISPRi-dCas9 System for Archaea and Its Use to Examine Gene Function during Nitrogen Fixation by Methanosarcina acetivorans. Applied and Environmental Microbiology, 86(21). https://doi.org/10.1128/AEM.01402-20

Edraki, A., Mir, A., Ibraheim, R., Gainetdinov, I., Yoon, Y., Song, C., Cao, Y., Gallant, J., Xue, W., Rivera-Pérez, J. A., & Sontheimer, E. J. (2019). A Compact, High-Accuracy Cas9 with a Dinucleotide PAM for in Vivo Genome Editing. Molecular Cell, 73(4), 714-726.e4. https://doi.org/10.1016/j.molcel.2018.12.003

Esquivel Álvarez, A., Quesada Morales, E., Villegas Calero, M. P., Matarrita Brenes, D., Rojas Salas, M. F., Madrigal Redondo, G., Chavarría Rojas, M., & Baltodano Viales, E. (2022). CRISPR-Cas Technology, the Tool of the Future. European Journal of Biology and Biotechnology, 3(2), 1-9. https://doi.org/10.24018/ejbio.2022.3.2.332

Galarza, V. O. (2019). Carbohidratos y proteínas en microalgas: potenciales alimentos funcionales. Brazilian Journal of Food Technology, 22. https://doi.org/10.1590/1981-6723.04319

Georgianna, D. R., Hannon, M. J., Marcuschi, M., Wu, S., Botsch, K., Lewis, A. J., ... & Mayfield, S. P. (2013). Production of recombinant enzymes in the marine alga Dunaliella tertiolecta. Algal Research, 2(1), 2-9.

Gómez Luna, L. M. (2007). Microalgas: aspectos ecologicos y biotecnologicos. Revista cubana de química, 19(2), 3.

Gómez, L., Orozco, M. I., Quiroga, C., Díaz, J. C., Huérfano, J., Díaz, L. E., Rodríguez, J., & Camacho K., J. E. (2019). Producción de Astaxantina y expresión de genes en Haematococcus pluvialis (Chlorophyceae, Volvocales) bajo condiciones de estrés por deficiencia de nitrógeno y alta irradiancia: Producción de astaxantina y expresión de genes en H. pluvialis. Mutis, 9(2), 7-24. https://doi.org/10.21789/22561498.1532

Greiner, A., Kelterborn, S., Evers, H., Kreimer, G., Sizova, I., & Hegemann, P. (2017). Targeting of Photoreceptor Genes in Chlamydomonas reinhardtii via Zinc-Finger Nucleases and CRISPR/Cas9. The Plant Cell, 29(10), 2498-2518. https://doi.org/10.1105/tpc.17.00659

Haft, D. H., Selengut, J., Mongodin, E. F., & Nelson, K. E. (2005). A Guild of 45 CRISPR-Associated (Cas) Protein Families and Multiple CRISPR/Cas Subtypes Exist in Prokaryotic Genomes. PLoS Computational Biology, 1(6), e60. https://doi.org/10.1371/journal.pcbi.0010060

Han, H. (2018). RNA Interference to Knock Down Gene Expression. Disease Gene Identification. Springer New York. https://doi.org/10.1007/978-1-4939-7471-9_16

Hopes, A., Nekrasov, V., Kamoun, S., & Mock, T. (2016). Editing of the urease gene by CRISPR-Cas in the diatom Thalassiosira pseudonana. Plant Methods, 12(1), 49. https://doi.org/10.1186/s13007-016-0148-0

Hu, L., Feng, S., Liang, G., Du, J., Li, A., & Niu, C. (2021). CRISPR/Cas9-induced β-carotene hydroxylase mutation in Dunaliella salina CCAP19/18. AMB Express, 11(1), 83. https://doi.org/10.1186/s13568-021-01242-4

Ishino, Y., Shinagawa, H., Makino, K., Amemura, M., & Nakata, A. (1987). Nucleotide sequence of the iap gene, responsible for alkaline phosphatase isozyme conversion in Escherichia coli, and identification of the gene product. Journal of Bacteriology, 169(12), 5429-5433. https://doi.org/10.1128/jb.169.12.5429-5433.1987

Jeon, S., Lim, J., Lee, H., Shin, S., Kang, N. K., Park, Y., Oh, H., Jeong, W., Jeong, B., & Chang, Y. K. (2017). Current status and perspectives of genome editing technology for microalgae. Biotechnology for Biofuels, 10(1), 267. https://doi.org/10.1186/s13068-017-0957-z

Jiang, W. Z., & Weeks, D. P. (2017). A gene-within-a-gene Cas9/sgRNA hybrid construct enables gene editing and gene replacement strategies in Chlamydomonas reinhardtii. Algal Research, 26(C), 474-480. https://doi.org/10.1016/j.algal.2017.04.001

Jiang, W., Brueggeman, A. J., Horken, K. M., Plucinak, T. M., & Weeks, D. P. (2014). Successful Transient Expression of Cas9 and Single Guide RNA Genes in Chlamydomonas reinhardtii. Eukaryotic Cell, 13(11), 1465-1469. https://doi.org/10.1128/EC.00213-14

Jiang, X., Liu, Y., Yin, X., Deng, Z., Zhang, S., Ma, C., & Wang, L. (2023). Efficient removal of chromium by a novel biochar-microalga complex: Mechanism and performance. Environmental Technology & Innovation, 31, 103156. https://doi.org/10.1016/j.eti.2023.103156

Kao, P., & Ng, I. (2017). CRISPRi mediated phosphoenolpyruvate carboxylase regulation to enhance the production of lipid in Chlamydomonas reinhardtii. Bioresource Technology, 245(Pt B), 1527-1537. https://doi.org/10.1016/j.biortech.2017.04.111

Kelterborn, S., Boehning, F., Sizova, I., Baidukova, O., Evers, H., & Hegemann, P. (2022). Gene Editing in Green Alga Chlamydomonas reinhardtii via CRISPR-Cas9 Ribonucleoproteins. Methods in molecular biology, 45-65. https://doi.org/10.1007/978-1-0716-1791-5_3

Kim, J., Chang, K. S., Lee, S., & Jin, E. (2021). Establishment of a Genome Editing Tool Using CRISPR-Cas9 in Chlorella vulgaris UTEX395. International Journal of Molecular Sciences, 22(2), 480. https://doi.org/10.3390/ijms22020480

Kim, M., Kim, S., Kim, Y., & Choi, T. (2023). Enhancement of Chlorella transformation efficacy by insert fragmentation. Algal Research (Amsterdam), 72, 103146. https://doi.org/10.1016/j.algal.2023.103146

Kleinstiver, B. P., Prew, M. S., Tsai, S. Q., Topkar, V. V., Nguyen, N. T., Zheng, Z., Gonzales, A. P. W., Li, Z., Peterson, R. T., Yeh, J. J., Aryee, M. J., & Joung, J. K. (2015). Engineered CRISPR-Cas9 nucleases with altered PAM specificities. Nature (London), 523(7561), 481-485. https://doi.org/10.1038/nature14592

Kolesnik, M. V., Fedorova, I., Karneyeva, K. A., Artamonova, D. N., & Severinov, K. V. (2021). Type III CRISPR-Cas Systems: Deciphering the Most Complex Prokaryotic Immune System. Biochemistry (Moscow), 86(10), 1301-1314. https://doi.org/10.1134/S0006297921100114

Kong, W., Shen, B., Lyu, H., Kong, J., Ma, J., Wang, Z., & Feng, S. (2021). Review on carbon dioxide fixation coupled with nutrients removal from wastewater by microalgae. Journal of Cleaner Production, 292, 125975. https://doi.org/10.1016/j.jclepro.2021.125975

Koonin, E. V., Makarova, K. S., & Zhang, F. (2017). Diversity, classification and evolution of CRISPR-Cas systems. Current Opinion in Microbiology, 37, 67-78. https://doi.org/10.1016/j.mib.2017.05.008

Kühn, R. (2009). Gene knockout protocols (2. ed. ed.). Humana Press.

Lakhawat, S. S., Malik, N., Kumar, V., Kumar, S., & Sharma, P. K. (2022). Implications of CRISPR-Cas9 in Developing Next Generation Biofuel: A Mini-review. Current Protein & Peptide Science, 23(9), 574-584. https://doi.org/10.2174/1389203723666220907110310

Le-Feuvre, R., Moraga-Suazo, P., Gonzalez, J., Martin, S. S., Henríquez, V., Donoso, A., & Agurto-Muñoz, C. (2020). Biotechnology applied to Haematococcus pluvialis Fotow: challenges and prospects for the enhancement of astaxanthin accumulation. Journal of Applied Phycology, 32, 3831-3852.

Li, C., Chu, W., Gill, R. A., Sang, S., Shi, Y., Hu, X., Yang, Y., Zaman, Q. U., & Zhang, B. (2022). Computational Tools and Resources for CRISPR/Cas Genome Editing. Genomics, Proteomics & Bioinformatics. https://doi.org/10.1016/j.gpb.2022.02.006

Li, Y., Xu, H., Han, F., Mu, J., Chen, D., Feng, B., & Zeng, H. (2015). Regulation of lipid metabolism in the green microalga Chlorella protothecoides by heterotrophy–photoinduction cultivation regime. Bioresource Technology, 192, 781-791. https://doi.org/10.1016/j.biortech.2014.07.028

Liao, C., & Beisel, C. L. (2021). The tracrRNA in CRISPR Biology and Technologies. Annual Review of Genetics, 55(1), 161-181. https://doi.org/10.1146/annurev-genet-071719-022559

Lin, W., & Ng, I. (2020). Development of CRISPR/Cas9 system in Chlorella vulgaris FSP-E to enhance lipid accumulation. Enzyme and Microbial Technology, 133, 109458. https://doi.org/10.1016/j.enzmictec.2019.109458

Liu, M., Rehman, S., Tang, X., Gu, K., Fan, Q., Chen, D., & Ma, W. (2019). Methodologies for Improving HDR Efficiency. Frontiers in Genetics, 9, 691. https://doi.org/10.3389/fgene.2018.00691

Luo, Q., Zou, X., Wang, C., Li, Y., & Hu, Z. (2021). The roles of Cullins E3 ubiquitin ligases in the lipid biosynthesis of the green microalgae Chlamydomonas reinhardtii. International Journal of Molecular Sciences, 22(9), 4695. https://doi.org/10.3390/IJMS22094695

Machado Sierra, E., Serrano, M. C., Manares, A., Guerra, A., & Aranguren Díaz, Y. (2021). Microalgae: Potential for Bioeconomy in Food Systems. Applied Sciences, 11(23), 11316. https://doi.org/10.3390/app112311316

Makarova, K. S., Grishin, N. V., Shabalina, S. A., Wolf, Y. I., & Koonin, E. V. (2006). A putative RNA-interference-based immune system in prokaryotes: computational analysis of the predicted enzymatic machinery, functional analogies with eukaryotic RNAi, and hypothetical mechanisms of action. Biology Direct, 1(1), 7. https://doi.org/10.1186/1745-6150-1-7

Makarova, K. S., Wolf, Y. I., Iranzo, J., Shmakov, S. A., Alkhnbashi, O. S., Brouns, S. J. J., Charpentier, E., Cheng, D., Haft, D. H., Horvath, P., Moineau, S., Mojica, F. J. M., Scott, D., Shah, S. A., Siksnys, V., Terns, M. P., Venclovas, Č, White, M. F., Yakunin, A. F., . . . Koonin, E. V. (2020). Evolutionary classification of CRISPR–Cas systems: a burst of class 2 and derived variants. Nature Reviews. Microbiology, 18(2), 67-83. https://doi.org/10.1038/s41579-019-0299-x

Mendoza, H., Jara, A. D. L., Freijanes, K., Cormona, L., Ramos, A. A., Duarte, V. D. S., et al. (2008). Characterization of Dunaliella salina strains by flow cytometry: A new approach to select carotenoid hyperproducing strains. Electronic Journal of Biotechnology, 11(4), 5–6. https ://doi.org/10.2225/vol11 –issue 4-fullt ext-2.

Musunuru, K., Chadwick, A. C., Mizoguchi, T., Garcia, S. P., DeNizio, J. E., Reiss, C. W., Wang, K., Iyer, S., Dutta, C., Clendaniel, V., Amaonye, M., Beach, A., Berth, K., Biswas, S., Braun, M. C., Chen, H., Colace, T. V., Ganey, J. D., Gangopadhyay, S. A., . . . Kathiresan, S. (2021). In vivo CRISPR base editing of PCSK9 durably lowers cholesterol in primates. Nature (London), 593(7859), 429-434. https://doi.org/10.1038/s41586-021-03534-y

Ng, I., Keskin, B. B., & Tan, S. (2020). A Critical Review of Genome Editing and Synthetic Biology Applications in Metabolic Engineering of Microalgae and Cyanobacteria. Biotechnology Journal, 15(8), e1900228-n/a. https://doi.org/10.1002/biot.201900228

Nino Castillo, C. M., Rodriguez Rivera, F. C., Diaz, L. E., & Lancheros Diaz, A. G. (2017). Evaluación de las condiciones de crecimiento celular para la producción de astaxantina a patir de la microalga Haematococcus pluvialis. Nova: Publicación Científica En Ciencias Biomédicas, 15(28), 19-31. https://doi.org/10.22490/24629448.2073

Nymark, M., Sharma, A. K., Sparstad, T., Bones, A. M., & Winge, P. (2016). A CRISPR/Cas9 system adapted for gene editing in marine algae. Scientific Reports, 6(1), 24951. https://doi.org/10.1038/srep24951

Ohlrogge, J., & Browse, J. (1995). Lipid biosynthesis. The plant cell, 7(7), 957.

Oslan, S. N. H., Shoparwe, N. F., Yusoff, A. H., Rahim, A. A., Chang, C. S., Tan, J. S., Oslan, S. N., Arumugam, K., Ariff, A. B., Sulaiman, A. Z., & Mohamed, M. S. (2021). A Review on Haematococcus pluvialis Bioprocess Optimization of Green and Red Stage Culture Conditions for the Production of Natural Astaxanthin. Biomolecules (Basel, Switzerland), 11(2), 256. https://doi.org/10.3390/biom11020256


Patel, V. K., Soni, N., Prasad, V., Sapre, A., Dasgupta, S., & Bhadra, B. (2019). CRISPR–Cas9 System for Genome Engineering of Photosynthetic Microalgae. Molecular Biotechnology, 61(8), 541-561. https://doi.org/10.1007/s12033-019-00185-3

Porta Helena. (2021). Conceptos básicos y avances de la transformación
genética de las microalgas, 25(5), 127-139.

Ren, Y., Deng, J., Huang, J., Wu, Z., Yi, L., Bi, Y., & Chen, F. (2021). Using green alga Haematococcus pluvialis for astaxanthin and lipid co-production: Advances and outlook. Bioresource Technology, 340, 125736. https://doi.org/10.1016/j.biortech.2021.125736

Safi, C., Zebib, B., Merah, O., Pontalier, P., & Vaca-Garcia, C. (2014). Morphology, composition, production, processing and applications of Chlorella vulgaris: A review. Renewable & Sustainable Energy Reviews, 35, 265-278. https://doi.org/10.1016/j.rser.2014.04.007

Schüler, L., Greque de Morais, E., Trovão, M., Machado, A., Carvalho, B., Carneiro, M., Maia, I., Soares, M., Duarte, P., Barros, A., Pereira, H., Silva, J., & Varela, J. (2020). Isolation and Characterization of Novel Chlorella Vulgaris Mutants with Low Chlorophyll and Improved Protein Contents for Food Applications. Frontiers in Bioengineering and Biotechnology, 8, 469. https://doi.org/10.3389/fbioe.2020.00469

Shah, M. M. R., Liang, Y., Cheng, J. J., & Daroch, M. (2016). Astaxanthin-Producing Green Microalga Haematococcus pluvialis: From Single Cell to High Value Commercial Products. Frontiers in Plant Science, 7, 531. https://doi.org/10.3389/fpls.2016.00531

Shams, F., Bayat, H., Mohammadian, O., Mahboudi, S., Vahidnezhad, H., Soosanabadi, M., & Rahimpour, A. (2022). Advance trends in targeting homology-directed repair for accurate gene editing: An inclusive review of small molecules and modified CRISPR-Cas9 systems. BioImpacts: BI, 12(4), 371-391. https://doi.org/10.34172/bi.2022.23871

Sharma, P. K., Saharia, M., Srivstava, R., Kumar, S., & Sahoo, L. (2018). Tailoring Microalgae for Efficient Biofuel Production. Frontiers in Marine Science, 5. https://doi.org/10.3389/fmars.2018.00382

Shin, S., Lim, J., Koh, H. G., Kim, E. K., Kang, N. K., Jeon, S., Kwon, S., Shin, W., Lee, B., Hwangbo, K., Kim, J., Ye, S. H., Yun, J., Seo, H., Oh, H., Kim, K., Kim, J., Jeong, W., Chang, Y. K., & Jeong, B. (2016). CRISPR/Cas9-induced knockout and knock-in mutations in Chlamydomonas reinhardtii. Scientific Reports, 6(1), 27810. https://doi.org/10.1038/srep27810

Shin, Y. S., Jeong, J., Nguyen, T. H. T., Kim, J. Y. H., Jin, E., & Sim, S. J. (2019). Targeted knockout of phospholipase A2 to increase lipid productivity in Chlamydomonas reinhardtii for biodiesel production. Bioresource Technology, 271, 368-374. https://doi.org/10.1016/j.biortech.2018.09.121

Sledzinski, P., Dabrowska, M., Nowaczyk, M., & Olejniczak, M. (2021). Paving the way towards precise and safe CRISPR genome editing. Biotechnology Advances, 49, 107737.

Spicer, A., & Molnar, A. (2018). Gene Editing of Microalgae: Scientific Progress and Regulatory Challenges in Europe. Biology (Basel, Switzerland), 7(1), 21. https://doi.org/10.3390/biology7010021

Tian, P., Wang, J., Shen, X., Rey, J. F., Yuan, Q., & Yan, Y. (2017). Fundamental CRISPR-Cas9 tools and current applications in microbial systems. Synthetic and Systems Biotechnology, 2(3), 219-225. https://doi.org/10.1016/j.synbio.2017.08.006

Trovão, M., Schüler, L. M., Machado, A., Bombo, G., Navalho, S., Barros, A., Pereira, H., Silva, J., Freitas, F., & Varela, J. (2022). Random Mutagenesis as a Promising Tool for Microalgal Strain Improvement towards Industrial Production. Marine Drugs, 20(7), 440. https://doi.org/10.3390/md20070440

Uddin, F., Rudin, C. M., & Sen, T. (2020). CRISPR Gene Therapy: Applications, Limitations, and Implications for the Future. Frontiers in Oncology, 10, 1387. https://doi.org/10.3389/fonc.2020.01387

Van der Oost, J., Jore, M. M., Westra, E. R., Lundgren, M., & Brouns, S. J. J. (2009). CRISPR-based adaptive and heritable immunity in prokaryotes. Trends in Biochemical Sciences (Amsterdam. Regular Ed.), 34(8), 401-407. https://doi.org/10.1016/j.tibs.2009.05.002

Velmurugan, A., & Muthukaliannan, G. K. (2022). Genetic manipulation for carotenoid production in microalgae an overview. Current Research in Biotechnology, 4, 221-228.

Vidhyavathi, R., Venkatachalam, L., Sarada, R., & Ravishankar, G. A. (2008). Regulation of carotenoid biosynthetic genes expression and carotenoid accumulation in the green alga Haematococcus pluvialis under nutrient stress conditions. Journal of experimental botany, 59(6), 1409-1418.

Wang, Q., Lu, Y., Xin, Y., Wei, L., Huang, S., & Xu, J. (2016). Genome editing of model oleaginous microalgae Nannochloropsis spp. by CRISPR/Cas9. The Plant Journal: For Cell and Molecular Biology, 88(6), 1071-1081. https://doi.org/10.1111/tpj.13307

Waissman-Levy, N., Leu, S., Khozin-Goldberg, I., & Boussiba, S. (2019). Manipulation of trophic capacities in Haematococcus pluvialis enables low-light mediated growth on glucose and astaxanthin formation in the dark. Algal Research, 40, 101497.

Wei, L., Xin, Y., Wang, Q., Yang, J., Hu, H., & Xu, J. (2017). RNAi‐based targeted gene knockdown in the model oleaginous microalgae Nannochloropsis oceanica. The Plant Journal: For Cell and Molecular Biology, 89(6), 1236-1250. https://doi.org/10.1111/tpj.13411

Wu, K., Ying, K., Zhou, J., Liu, D., Liu, L., Tao, Y., Hanotu, J., Zhu, X., & Cai, Z. (2021). Optimizing the growth of Haematococcus pluvialis based on a novel microbubble-driven photobioreactor. iScience, 24(12), 103461. https://doi.org/10.1016/j.isci.2021.103461

Xu, H., Xiao, T., Chen, C., Li, W., Meyer, C. A., Wu, Q., Wu, D., Cong, L., Zhang, F., Liu, J. S., Brown, M., & Liu, X. S. (2015). Sequence determinants of improved CRISPR sgRNA design. Genome Research, 25(8), 1147-1157. https://doi.org/10.1101/gr.191452.115

Xu, P., Li, J., Qian, J., Wang, B., Liu, J., Xu, R., Chen, P., & Zhou, W. (2023). Recent advances in CO2 fixation by microalgae and its potential contribution to carbon neutrality. Chemosphere (Oxford), 319, 137987. https://doi.org/10.1016/j.chemosphere.2023.137987

Yoshimitsu, Y., Abe, J., & Harayama, S. (2018). Cas9-guide RNA ribonucleoprotein-induced genome editing in the industrial green alga Coccomyxa sp. strain KJ. Biotechnology for Biofuels, 11(1), 326. https://doi.org/10.1186/s13068-018-1327-1

Yue Yang, Jeong Min Seo, Nguyen, A., Pham, T. X., Hea Jin Park, Park, Y., Kim, B., Bruno, R. S., & Jiyoung Lee. (2011). Astaxanthin-Rich Extract from the Green Alga Haematococcus pluvialis Lowers Plasma Lipid Concentrations and Enhances Antioxidant Defense in Apolipoprotein E Knockout Mice. The Journal of Nutrition, 141(9), 1611-1617. https://doi.org/10.3945/jn.111.142109

Zambrano, J., García-Encina, P. A., Jiménez, J. J., Ciardi, M., Bolado-Rodríguez, S., & Irusta-Mata, R. (2023). Removal of veterinary antibiotics in swine manure wastewater using microalgae-bacteria consortia in a pilot scale photobioreactor. Environmental Technology & Innovation, 103190. https://doi.org/10.1016/j.eti.2023.103190

Zetsche, B., Gootenberg, J., Abudayyeh, O., Slaymaker, I., Makarova, K., Essletzbichler, P., Volz, S., Joung, J., van der Oost, J., Regev, A., Koonin, E., & Zhang, F. (2015). Cpf1 Is a Single RNA-Guided Endonuclease of a Class 2 CRISPR-Cas System. Cell, 163(3), 759-771. https://doi.org/10.1016/j.cell.2015.09.038

Zhang, S., Guo, F., Yan, W., Dai, Z., Dong, W., Zhou, J., Zhang, W., Xin, F., & Jiang, M. (2020). Recent Advances of CRISPR/Cas9-Based Genetic Engineering and Transcriptional Regulation in Industrial Biology. Frontiers in Bioengineering and Biotechnology, 7, 459. https://doi.org/10.3389/fbioe.2019.00459

Zhang, Y., Jiang, J., Shi, T., Sun, X., Zhao, Q., Huang, H., & Ren, L. (2019). Application of the CRISPR/Cas system for genome editing in microalgae. Applied Microbiology and Biotechnology, 103(8), 3239-3248. https://doi.org/10.1007/s00253-019-09726-x

Zhu, Y. (2022). Advances in CRISPR/Cas9. BioMed Research International, 1-13. https://doi.org/10.1155/2022/9978571

Citado por