...it grows fast to boot. The yeast strains S. cerevisiae and S. pombe have dominated the research scene....the most frequently used yeast strains: S. pombe (fission) and S. cerevisiae (budding). Yeast make for ...probably wondering if the “biotech yeast” is S. pombe or S. cerevisiae. Actually, it’s neither. The most...phaffii and other yeast strains.     S. cerevisiae S. pombe K. phaffii Growth properties ...glycerol). K. phaffii is only distantly related to S. pombe and S. cerevisiae; evolutionarily it evolved much... K. phaffii to work   The yeast giants - S. cerevisiae and S. pombe - aren’t going anywhere anytime soon...Amenable to genetic manipulation The first eukaryote (S. cerevisiae) to have its genome fully sequenced There...

...source. S. cerevisiae  no no   LEU2 L-leucine no S. cerevisiae yes - This can complement leu1- S. pombe...L-hisitidine no S. cerevisiae  no yes   URA3 pyrimidine (uracil) yes - Grow with 5-FOA. S. cerevisiae yes... ura4+. leu1+ L-leucine   S. pombe  no no   ade6+ purine (adenine)   S. pombe  no no   Considerations...are included (reviewed here). Plasmids for use in S. pombe, on the other hand, do not require a well defined...sequence) dictate the replication of these vectors. S. pombe plasmids oftentimes utilize an ARS to aid in... yes - This can complement ura4- S. pombe, but the complementation is weak. yes   LYS2 L-lysine yes ... yes   TRP1 L-tryptophan yes - Grow with 5-FAA. S. cerevisiae no  no TRP1 alters some yeast phenotypes...

...Osuka, S., Isomura, K., Kajimoto, S., Komori, T., Nishimasu, H., Shima, T., Nureki, O., & Uemura, S. (2018... J. P. K., Liu, M.-S., Hibshman, G. N., Dangerfield, T. L., Jung, K., McCool, R. S., Johnson, K. A., &...Nishimasu, H., Ran, F. A., Hsu, P. D., Konermann, S., Shehata, S. I., Dohmae, N., Ishitani, R., Zhang, F., &...://doi.org/10.1038/nature15544 Sternberg, S. H., Redding, S., Jinek, M., Greene, E. C., & Doudna, J. A....   Figure 1: A cartoon depiction of Cas9’s two major lobes, REC and NUC, and their subdomains...pyogenes.   Figure 2:  Crystal structure of S. pyogenes Cas9 in the apo state (PDB ID 4CMP) with...can offer more information on these regions.   Cas9’s transition from weak, random binding to precision ...

...rearrangements in the S. cerevisiae lineage (7). In our class research project, students transform S. cerevisiae...plasmids carrying MET/Met genes from either S. pombe or S. cerevisiae. Students then determine if expression...shown that most, but not all, S. pombe Met genes complement the corresponding S. cerevisiae met deficiencies...strains is available (6). Methionine synthesis in S. cerevisiae (Figure 1) occurs through a well-characterized..., have the same function as their counterparts in S. cerevisiae. The two yeasts are thought to have diverged... the plasmids in either E. coli (ori and ampR) or S. cerevisiae (2µm ori and URA3). The presence of the...successfully transformed with the plasmids, because the S. cerevisiae deletion strains are ∆ura3 mutants and...

...Addgene. Kim D, Kim J, Hur JK, Been KW, Yoon S, Kim J-S (2016) Genome-wide analysis reveals specificities...Cho H-Y, Song DW, Lee KJ, Jung MH, Kim S, Kim JH, Kim JH, Kim J-S (2017) In vivo genome editing with a ...sequences While PAM sequences for the commonly used S. pyogenes Cas9 (3'-NGG) are abundant throughout the...to circumvent this limitation: 1) the use of novel S. pyogenes Cas9 variants with varying PAM sequences... of Cas9 homologs derived from species other than S. pyogenes, and 3) the use of non-Cas9 enzymes. (For...site, check out this video from IGI!) Synthetic S. pyogenes Cas9s with novel PAM recognition In 2015...selection screens in bacteria to identify mutants of S. pyogenes Cas9 that were able to cleave target DNA...

...and happy optimizing!  Recommended Reading Issa, S. S., Shaimardanova, A. A., Solovyeva, V. V., & Rizvanov...Resources References Aschauer, D. F., Kreuz, S., & Rumpel, S. (2013). Analysis of Transduction Efficiency...10.1089/hum.2009.169 Dudek, A. M., Pillay, S., Puschnik, A. S., Nagamine, C. M., Cheng, F., Qiu, J., Carette... https://doi.org/10.1038/s41434-019-0075-6 Issa, S. S., Shaimardanova, A. A., Solovyeva, V. V., & Rizvanov...10.1371/journal.pone.0076310 Damdindorj, L., Karnan, S., Ota, A., Hossain, E., Konishi, Y., Hosokawa, Y.,...Fong, D. M., Mouravlev, A., Young, D., & O’Carroll, S. J. (2019). Astrocyte-selective AAV gene therapy through...cells12050785 Kanaan, N. M., Sellnow, R. C., Boye, S. L., Coberly, B., Bennett, A., Agbandje-McKenna, M...

...depletion assay to discover FnCpf1’s PAM sequence requirements. Cpf1’s preferred PAM is 5’-TTN, differing...RuvC-like endonuclease domain, but they lack Cas9’s second HNH endonuclease domain. Cpf1 cleaves DNA in... RuvC-like endonuclease domain, but it lacks Cas9’s other HNH endonuclease domain, indicating that Cpf1... nuclease could potentially overcome some of Cas9’s shortcomings - namely its blunt double stranded cleavage...nucleotides in length, about the same size as Cas9’s, but with the direct repeat preceding the spacer rather...crRNA is also much simpler in structure than Cas9’s; only a short stem-loop structure in the direct repeat... added another option to the CRISPR toolbox. Cpf1’s staggered cleavage pattern opens up the possibility...

...integrate protein tags into the genomes of S. cerevisiae and S. pombe. The protocol is surprisingly simple...Yeast-optimized fluorophores for imaging Lee S, et al. (2013) PLoS ONE 8(7): e67902. Many imaging...al.(1) assessed many of these fluorescent tags in S. cerevisiae, looking at their performance in categories...plasmids for a wide variety of genome modifications in S. Pombe, including full and partial gene deletion, ...describe a complimentary set of plasmids for use in S. cerevisiae, with the additional benefit of multiple...system that isn't mentioned here? References: Lee S, Lim WA, Thorn KS. PLoS One. 2013 Jul 2;8(7):e67902...

...Erdmann, R. S., Baguley, S. W., Richens, J. H., Wissner, R. F., Xi, Z., Allgeyer, E. S., Zhong, S., Thompson...F., Kromann, E. B., Allgeyer, E. S., Erdmann, R. S., Wood Baguley, S., Sirinakis, G., Schepartz, A., Baddeley...doi.org/10.1016/j.chembiol.2019.01.003 Gallagher, S. S., Sable, J. E., Sheetz, M. P., & Cornish, V. W. ... R., Bewersdorf, J., Rothman, J. E., Johnston, D. S., Schepartz, A., & Toomre, D. (2019). Labeling Strategies...10.1016/j.chembiol.2008.01.007 Griffin, B. A., Adams, S. R., & Tsien, R. Y. (1998). Specific Covalent Labeling...doi.org/10.1021/ar200099f Keppler, A., Gendreizig, S., Gronemeyer, T., Pick, H., Vogel, H., & Johnsson,...s41467-023-40788-8 Ulrichs, H., Gaska, I., & Shekhar, S. (2023). Multicomponent regulation of actin barbed...

...Schüchner, et al.’s results mean the end of 9E10’s time in the saga of the Myc tag? Hardly! 9E10’s accessibility...generated in an attempt to overcome some of Myc1-9E10’s shortcomings. Wait, 9E10 isn’t the Mary Poppins of... acids neighboring the Myc epitope influence 9E10’s affinity for the epitope, a phenomenon known as sequence...https://doi.org/10.1128/mcb.5.12.3610-3616.1985 Munro S, Pelham HR (1984) Use of peptide tagging to detect...doi.org/10.1002/j.1460-2075.1984.tb02263.x  Munro S, Pelham HRB (1986) An hsp70-like protein in the ER...https://doi.org/10.1016/0092-8674(86)90746-4 Munro S, Pelham HRB (1987) A C-terminal signal prevents secretion...//doi.org/10.1016/0092-8674(87)90086-9  Schüchner S, Behm C, Mudrak I, Ogris E (2020) The Myc tag monoclonal...

... Kumar, S., Chen, X., Goertsen, D., Coughlin, G. M., Hori, A. M., Chuapoco, M. R., Otis, T. S., Miles,...Ichiki, T., Mazmanian, S. K., Osborne, P. B., Keast, J. R., Miller, C. T., Fox, A. S., Chiu, I. M., & Gradinaru... Bindu, D. S., Zhang, M., Taskin, N., Goertsen, D., Shay, T. F., Sullivan, E. E., Huang, S.-F., Ravindra...B. P., Kumar, S. R., Chan, K. Y., Banerjee, A., Wu, W.-L., Yang, B., Huber, N., Pasca, S. P., & Gradinaru...., Ng, C., Nammour, J., Maiz, D., Dujardin, S., Ramirez, S. H., Hudry, E., & Maguire, C. A. (2019). Selection...Liguore, W. A., Domire, J. S., Button, D., Wang, Y., Dufour, B. D., Srinivasan, S., & McBride, J. L. (2019...Stanton, A., King, E. M., Ye, S., Tellez, L., Krunnfusz, A., Tavakoli, S., Widrick, J. J., Messemer, K...

...viability of your cells. Enrich S-phase cells HR operates predominantly in S and G2 phase of the cell cycle....1016/j.jbiotec.2019.11.010 Wright, D. W., Shah, S. S., Heyer, W. D. Homologous recombination and the ...2018). 1 10.1074/jbc.TM118.000372 Yang, H., Ren, S., Yu, S., Pan, H., et al. Methods favoring homology-directed...regulatory proteins responsible for the transition out of S phase, so cells stay in an HDR-promoted cycle longer...., 21(18): 6461 (2020). 10.3390/ijms21186461 Lin, S., Staahl, B. T., Alla, R. K., Doudna, J. A. Enhanced...01836-2 Charpentier, M., Khedher, A. H. Y., Menerot, S., Brion, A., et al. CtIP fusion to Cas9 enhances transgene...

...10.1038/nn.2495 Marshel, J. H., Kim, Y. S., Machado, T. A., Quirin, S., Benson, B., Kadmon, J., Raja, C., ..., J. C., McKnight, D. J., Yoshizawa, S., Kato, H. E., Ganguli, S., & Deisseroth, K. (2019). Cortical layer–specific...following factors will help you determine which one(s) are candidates for your experiments.  Wavelengths....1007/978-1-0716-0755-8_2 Lehtinen, K., Nokia, M. S., & Takala, H. (2022). Red Light Optogenetics in Neuroscience...://doi.org/10.1007/978-1-0716-0755-8_2 Boyden, E. S., Zhang, F., Bamberg, E., Nagel, G., & Deisseroth,...Gunaydin, L. A., Yizhar, O., Berndt, A., Sohal, V. S., Deisseroth, K., & Hegemann, P. (2010). Ultrafast....1808 Nagel, G., Szellas, T., Huhn, W., Kateriya, S., Adeishvili, N., Berthold, P., Ollig, D., Hegemann...

...-excited GFP (vex) Lineage-Cre; Rosa26-Cas9 S-CHIME Knocks out pairs of genes sequentially pXPR...hindering studies post embryogenesis. Using C-CHIME and S-CHIME, we discovered that double knockout of PTPN1...References LaFleur, M. W., Lemmen, A. M., Streeter, I. S. L., Nguyen, T. H., Milling, L. E., Derosia, N. M...., Z. M., Gillis, J. E., Tjokrosurjo, Q., Markson, S. C., Huang, A. Y., Anekal, P. V., Montero Llopis, ...Coxe, M. A., Yates, K. B., Trombley, J. D., Weiss, S. A., Brown, F. D., Gillis, J. E., Coxe, D. J., Doench...

... et al for more examples of GeoCas9’s editing activity in HEK293’s. GeoCas9 stability in human plasma ...SpCas9 can tolerate. Harrington et al tested GeoCas9’s love of heat in two ways. The first set of experiments... mammalian cells. Harrington et al tested GeoCas9’s editing skills by transfecting HEK293 cells with GeoCas9...organisms at lower than optimal temperatures. GeoCas9’s thermostability could also be used with in vitro molecular...

...., Bull, R. L., Garcia, K. Y., Goodrich, J. S., Lovett, S. P., Parker, K., Radune, D., Hatada, A., Pan...Reyes-Sandoval, A., Harris-McCoy, K. C., Hensley, S. E., Zhou, D., Lin, S. W., Bian, A., Zhi, Q. X., Iparraguirre...Scaria, A., Hermiston, T. W., Ryerse, J. S., Wold, L. J., & Wold, W. S. (1996). The adenovirus death protein...Implications for gene therapy. Proc Natl Acad Sci U S A, 95(19), 11377. https://doi.org/10.1073/PNAS.95.19.11377.../JVI.70.4.2296-2306.1996. PMID: 8642656. Wold, W. S. M., & Toth, K. (2013). Adenovirus Vectors for Gene...

...team replaced S. pyogenes Cas9 in the ABE complex with PAM-variant Cas9 proteins: the S. pyogenes NG-Cas9... TadA and TadA7.10, they designed one set of ABE8’s similarly (ABE8.x-d). In order to assess the effect...created ABE8 constructs using a catalytically “dead” S. pyogenes Cas9 (dC9-ABE8.x-m/d). The dC9-ABE8.x-m/...scope of adenine base editors The most commonly used S. pyogenes Cas9 requires the target site in the genomic...-Cas9 (PAM: NG) to create NG-ABE8.xm/d and the S. aureus Cas9 (PAM: NNGRRT) to create Sa-ABE8.x-m/d. With...NM, Barrera LA, Born DA, Edwards A, Gehrke JM, Lee S-J, Liquori AJ, Murray R, Packer MS, Rinaldi C, Slaymaker...

...C-Y, Gootenberg JS, Konermann S, Trevino AE, Scott DA, Inoue A, Matoba S, Zhang Y, Zhang F (2013) Double.... https://doi.org/10.1038/ncomms5286 Yan M-Y, Li S-S, Ding X-Y, Guo X-P, Jin Q, Sun Y-C (2020) A CRISPR-Assisted...Piganeau M, Renouf B, Renaud J-B, Sallmyr A, Ruis B, Oh S, Tomkinson AE, Hendrickson EA, Giovannangeli C, Jasin...Pryor JM, Strom CN, Mieczkowski P, Burkhalter MD, Oh S, Qaqish BF, Moore DT, Hendrickson EA, Ramsden DA (...

...Levesque, S., Jing, Y., Simpson, D., Zhao, A., Li, H., Yan, W., Goudy, L., Schmidt, R., Solley, S. C., Gilbert...domains derived from the E. coli Ec48 retron RT and the S. pombe Tf1 retrotransposon RT, respectively. Their...Newby, G. A., Sahin, M., Osborn, M. J., Weissman, J. S., Adamson, B., & Liu, D. R. (2021). Enhanced prime.../10.1038/s41587-021-01025-z Doman, J. L., Pandey, S., Neugebauer, M. E., An, M., Davis, J. R., Randolph...Raguram, A., Richter, M. F., Everette, K. A., Banskota, S., Tian, K., Tao, Y. A., Tolar, J., Osborn, M. J., .../doi.org/10.1016/j.cell.2023.07.039 Lin, Q., Jin, S., Zong, Y., Yu, H., Zhu, Z., Liu, G., Kou, L., Wang...021-00868-w Nelson, J. W., Randolph, P. B., Shen, S. P., Everette, K. A., Chen, P. J., Anzalone, A. V....

....1101/2023.03.20.533580 Goodwin, M. S., Croft, C. L., Futch, H. S., Ryu, D., Ceballos-Diaz, C., Liu, X...Truong, V., Rostonics, E., Hernandez, A., Boye, S. L., Boye, S. E., Levites, Y., … Golde, T. E. (2020). Utilizing...producer cells for further processing. What component(s) you need to collect will depend on the serotype, ....1089/hum.2020.120 Lock, M., Alvira, M. R., Chen, S.-J., & Wilson, J. M. (2014). Absolute determination...doi.org/10.1089/hgtb.2013.131 Lock, M., McGorray, S., Auricchio, A., Ayuso, E., Beecham, E. J., Blouin-Tavel...