...Haoyi Qiu.   He Tian: Directed evolution of biosensors He Tian (goes by “Tian”) is a postdoc at Harvard...Brandán Pedre Pérez Elizabeth Hubbard Haoyi Qiu He Tian Every two years, we seek applications for awards...engineers genetically encoded voltage indicators. Tian developed a video-based high-throughput screen for... to identify the mutants for further enrichment. Tian notes that this platform can be used to engineer...types of biosensors.   This screening method allowed Tian to optimize a red‑shifted archaerhodopsin‑derived...neurotransmitter, calcium, and membrane voltage,” says Tian. Figure 5: Citrine-tagged voltage indicator...indicator expressed in mouse cortex. Image from He Tian. Congrats again to all the recipients! ...

...pAAV-CAG-RdLight1 AAV9 Biosensor Tian New viral vector pAAV-hSynapsin1-RdLight1 AAV9 Biosensor Tian New viral vector...deposited by the St-Pierre lab. Also of interest is the Tian lab's pAAV_hSynapsin_psychLight2 (AAV9), which contains...vector pAAV_hSynapsin_psychLight2 AAV9 Biosensor Tian New viral vector pAAV.hSynapsin.SF-iGluSnFR.A184S...reports from other users. And don't forgot to use the link below to keep up with the latest in Addgene's viral...

...expression of dLight1.1 under synapsin promoter Lin Tian 111067-AAV1 pAAV-CAG-dLight1.1 To generate Adeno-Associated...for expression of dLight1.1 under CAG promoter Lin Tian 111067-AAV5 pAAV-CAG-dLight1.1 To generate Adeno-Associated...for expression of dLight1.1 under CAG promoter Lin Tian 111068-AAV1 pAAV-hSyn-dLight1.2 To generate Adeno-Associated...expression of dLight1.2 under synapsin promoter Lin Tian 111068-AAV5 pAAV-hSyn-dLight1.2 To generate Adeno-Associated...expression of dLight1.2 under synapsin promoter Lin Tian 111069-AAV1 pAAV-CAG-dLight1.2 To generate Adeno-Associated...for expression of dLight1.1 under CAG promoter Lin Tian 111261-AAV9 pAAV-hSynapsin1-axon-GCaMP6m For enriched... enriched expression of GCaMP6m in axons Lin Tian 111262-AAV1 pAAV-hSynapsin1-axon-GCaMP6s For enriched...

...Constitutive 1 Tian 112000 pAAV-hSynapsin1-axon-GCaMP6f Syn axon-GCaMP6f none Constitutive 1 Tian 112005 pAAV-hSynapsin1...5, 9 Tian 112010 pAAV-hSynapsin1-FLEx-axon-GCaMP6s Syn axon-GCaMP6s none Cre dependent 5, 9 Tian Calcium...Constitutive 1 Tian 111067 pAAV-CAG-dLight1.1 CAG dLight1.1 none Constitutive 1, 5 Tian 111068 pAAV-hSyn-dLight1.2...Constitutive 1, 5 Tian 125560 AAV-CAG-dLight1.3b CAG dLight1.3b none Constitutive 1, 5, 9 Tian 135762 pAAV-...none Constitutive 9 Tian 125707 pAAV-CAG-RdLight1 CAG RdLight1 none Constitutive 9 Tian 125708 pAAV-hSynapsin1...Constitutive 9 Tian 128486 pAAV-CAG-FLEx-iSeroSnFR CAG iSeroSnFR none Cre dependent 9 Tian 163909 pAAV_hSynapsin_psychLight2...axon-GCaMP6m Syn axon-GCaMP6m none Constitutive 9 Tian 131004 pAAV-hSyn-GCaMP 6m-p2A-ChRmine-Kv2.1-WPRE...

...Figure 2). The dLight1 dopamine sensor In 2018, Lin Tian’s lab has developed dLight1, a molecular tool to...  Using dLight1 sensors in experiments The Tian lab used dLight1 both in brain slices and freely behaving...Feelings of delight associated with the brain’s reward circuits are generally linked to dopamine (DA)...exhibit fast kinetics (10 ms on and 100 ms off), enabling detection of individual peaks of DA release.  ... neurons in live mice Report modulation of DA signaling during learning and reward consumption in live...

...serotype pAAV-CAG-FLEx-iSeroSnFR AAV9 Biosensors Lin Tian New viral prep pAAV-hSyn-GRAB-gDA3m AAV1 Biosensors...reports from other users. And don't forget to use the link below to keep up with the latest in Addgene's viral...

...goal of Elizabeth Unger’s postdoctoral work in Lin Tian’s lab at UC Davis is to generate a genetically ...recognize serotonin. Using a combination of computer modeling, directed evolution, site-saturation mutagenesis...

...10.1089/hgtb.2019.276. PMID: 31752513. Zhou, Q., Tian, W., Liu, C., Lian, Z., Dong, X., & Wu, X. (2017...the gene of interest. A single ITR consists of palindromic arms (A-A’, B-B’, and C-C’) that give the feature...-shape. The arrangement of the B-B’ and C-C’ palindromic sequences determines the orientation of the ITR...and flop orientation. The ITR contains three palindromic sequences (A-A’, B-B’, and C-C’ regions) and ...

.... 2018 Jun 29;360(6396). pii: science.aat4422. Lin Tian Dopamine Yellow or Red genetically encoded dopamine... vivo. Nat Methods. 2020 Nov;17(11):1147-1155. Lin Tian Endocannabinoid GRAB_eCB2.0/eCBmut biosensor for...Cell. 2020 Dec 12. pii: S0092-8674(20)31612-3. Lin Tian Serotonin Serotonin (5-HT) sensor psychLight Psychedelic-inspired... Cell. 2021 Apr 8. pii: S0092-8674(21)00374-3. Lin Tian Serotonin Cilia-targeted serotonin sensor GRAB-HTR6...In Vivo. Cell. 2016 Jun 30;166(1):245-57. Michael Lin Voltage Ultrafast fluorescent voltage sensor for ...voltage sensor. Nat Neurosci. 2014 Apr 22. Michael Lin Voltage Fluorescent voltage reporters Archon 1 and...high-light signalling mechanism. Nat Commun. 2017 Jun 29;8(1):49. Philip Mullineaux Insulin Sensor for...

...viral preps here AAV-CAG-dLight1.3b from the Lin Tian Lab is now available as a ready to use viral prep...24390935/ Hypercompact genome editor CRISPR-Casɸ By Melina Fan The Doudna Lab adds to the CRISPR-Cas toolbox...target footprint compared to current N-terminal linked SaCas9 ABE variants. It's also one of the smallest...

...Frontiers in molecular neuroscience. 2014 Nov 18;7:88. Tian L, Hires SA, Looger LL. Imaging neuronal activity...teeth. It is an enzyme co-factor, a crosslinking agent, a signaling ion, and an allosteric modulator. Because...are fluorescent proteins linked to calcium binding proteins (e.g., calmodulin or troponin C). Their fluorescent...aberrant binding of the biosensor to endogenous calmodulin. Multiplexing Are you combining your calcium ...pdb-top069609. Palmer AE, Tsien RY. Measuring calcium signaling using genetically targetable fluorescent indicators...

...read up on the timing of this approach carefully (Lin et al., eLife)! Use a Cas that facilitates HR What...Resources and References References Yu, Y., Guo, Y., Tian, Qigi, Lan, Y., et. al. An efficient gene knock-... Sci., 21(18): 6461 (2020). 10.3390/ijms21186461 Lin, S., Staahl, B. T., Alla, R. K., Doudna, J. A. Enhanced...breast and ovarian lines commonly used in lab. It is recommended you check your cell lines for HR-related ...road less traveled and generating a knock-in cell line instead of a knock-out? We’ve got you covered in...engineering The easiest way to make a knock-in cell line is to utilize the built-in repair pathway cells ...maximize HDR events, ensure cells are actively cycling through the cell cycle stages. If cells are overly...

...from recent months: Axon targeting GCaMP6f from Lin Tian’s lab is available in serotype AAV1. The GCaMP8...enhance axonal labeling and synaptophysin-tagged mRuby to enrich pre-synaptic labeling. Now available ...This approach can augment viral surveillance by enabling rapid assessment of antigenic drift.    Starr...immunoprecipitate the entire complex in three different cell lines. The Gräslund lab plans to continue their work by...photocleavable gRNAs within seconds. A CRISPR-based lineage tracing tool captures rare clones that represent...where gRNA expression is induced by anhydrous tetracycline. Three new anti-CRISPRs were identified that...

... A., Danner-Bocks, S., Sanghal, T., Yadanar, L., Tian, R., Ma, K., Palop, J. J., & Kampmann, M. (2023)... the gRNA and scaffold. If you don’t feel like doubling the amount of cloning, another option is to use...doi.org/10.1101/2023.06.13.544831 Santinha, A. J., Klingler, E., Kuhn, M., Farouni, R., Lagler, S., Kalamakis...Jabaudon, D., & Platt, R. J. (2023). Transcriptional linkage analysis with in vivo AAV-Perturb-seq. Nature, ...

... Central PMCID: PMC4927352. Janetopoulos, Chris, Tian Jin, and Peter Devreotes. "Receptor-mediated activation...Therefore, the localization of critical fusions (e.g. tubulin or connexin) has been used as a proxy for dimerization...mNeonGreen, mScarlet(-I), and mCherry. Keep the linker sequence short In the early days of GFP applications...protein. Thus, scientist began using relatively long linkers between the protein of interest and the fluorescent...small, non-aliphatic amino acids. However, long linkers increase the chance that a fusion protein is cleaved...victoria GFP sticks out and can be considered a linker (figure 1). In fact, for several FRET biosensors...Lifeact-mTurquoise2 (Plasmid #36201) or mScarlet-tubulin (Plasmid #85045). We often use restriction enzymes...

...0513-4 Chen JS, Ma E, Harrington LB, Da Costa M, Tian X, Palefsky JM, Doudna JA (2018) CRISPR-Cas12a target...specific nucleic acid detection is crucial for clinical diagnostics, genotyping, and biotechnological ...be used for an array of applications including clinical diagnostics (e.g. pathogen or virus detection)...points out that the protocol is not approved for clinical use at this point, they hope that the protocol...mixed population for an array of molecular and clinical diagnostic applications. The company Mammoth Biosciences...Livny J, Regev A, Koonin EV, Hung DT, Sabeti PC, Collins JJ, Zhang F (2017) Nucleic acid detection with ...Gootenberg JS, Abudayyeh OO, Kellner MJ, Joung J, Collins JJ, Zhang F (2018) Multiplexed and portable nucleic...

..., Richter, M. F., Everette, K. A., Banskota, S., Tian, K., Tao, Y. A., Tolar, J., Osborn, M. J., & Liu....e26. https://doi.org/10.1016/j.cell.2023.07.039 Lin, Q., Jin, S., Zong, Y., Yu, H., Zhu, Z., Liu, G.,...been identified in humans and cataloged in the ClinVar database. Previously developed genome editing methods... In addition to the numerous innovations that streamline the process of prime editing, the Liu lab contributed... 3′ end, the Liu lab appended RNA pseudoknots, calling this longer RNA an “engineered pegRNA” or epegRNA...long way to go before we see prime editors in the clinic. We look forward to following new advances in this...

...resolution. PLoS One. 2(3):e299. PMID 17375185 Kim B, Lin MZ. 2013. Optobiology: optical control of biological...A, Cho YK, Morimoto TK, Chuong AS, Carpenter EJ, Tian Z, et al. 2014. Independent optical excitation of...populations. Nat Methods. 11(3):338-46. PMID 24509633 Lin, J. 2011. A User’s Guide to Channelrhodopsin Variants...chloride pump from Haloarcula (Halobacterium) salinarum (strain Shark), used to silence neuronal activity...stable expression is achieved in a transgenic mouse line, such as for example the VGAT-ChR2 mouse, where ...and the light oxygen voltage (LOV) domain of FKF1 LINUS light-inducible nuclear localization signal based...local concentration near the cell surface, and enabling the ion channel to function. TULIPs Tunable light-inducible...

...1171. PMID: 23979020 Cong, L., Ran, F. A., Cox, D., Lin, S., Barretto, R., Habib, N., Hsu, P. D., Wu, X.,...Science , 339 (6121), 819–823. PMID: 23287718 Cui, Z., Tian, R., Huang, Z., Jin, Z., Li, L., Liu, J., Huang,...L., Sun, N., Zeina, C. M., Gao, X., Rees, H. A., Lin, Z., & Liu, D. R. (2018). Evolved Cas9 variants with..., Richter, M. F., Everette, K. A., Banskota, S., Tian, K., Tao, Y. A., Tolar, J., Osborn, M. J., & Liu...Molecular Sciencess . 25 (5), 2456. PMID: 38473704 Lin, S., Staahl, B. T., Alla, R. K., & Doudna, J. A. ..., S., Pruitt, B. W., Tuttle, M., Iyer, E. P. R., Lin, S., Kiani, S., Guzman, C. D., Wiegand, D. J., Ter-Ovanesyan...single-stranded DNA, unraveling DNA as it goes using its helicase properties, often called a reeling mechanism. ...

...1171. PMID: 23979020 Cong, L., Ran, F. A., Cox, D., Lin, S., Barretto, R., Habib, N., Hsu, P. D., Wu, X.,...Science , 339 (6121), 819–823. PMID: 23287718 Cui, Z., Tian, R., Huang, Z., Jin, Z., Li, L., Liu, J., Huang,...L., Sun, N., Zeina, C. M., Gao, X., Rees, H. A., Lin, Z., & Liu, D. R. (2018). Evolved Cas9 variants with..., Richter, M. F., Everette, K. A., Banskota, S., Tian, K., Tao, Y. A., Tolar, J., Osborn, M. J., & Liu...Molecular Sciencess . 25 (5), 2456. PMID: 38473704 Lin, S., Staahl, B. T., Alla, R. K., & Doudna, J. A. ..., S., Pruitt, B. W., Tuttle, M., Iyer, E. P. R., Lin, S., Kiani, S., Guzman, C. D., Wiegand, D. J., Ter-Ovanesyan...single-stranded DNA, unraveling DNA as it goes using its helicase properties, often called a reeling mechanism. ...