...capsids from serotypes 1, 2, and 5 display differential efficiency and cell tropism after delivery to ...indicates a virus containing the genome of serotype 2 packaged in the capsid from serotype 5. Use of these...neurons that are not efficiently transduced by AAV2/2, and is distributed more widely in the brain, indicating...PJ, Mandel RJ, Muzyczka N. Mol. Ther. 2004 Aug;10(2):302-17. PubMed . From virus evolution to vector revolution...persist in non-dividing cells for a period of years without damaging the host cell. In contrast, viral particles...through cell lysis. Episomal stability enables long-term transgene expression in non-dividing cells and is... the helper plasmid are transfected into HEK293 cells, which contain the adenovirus gene E1+, to produce...

....9 (parental) 46 +/- 0.7% AAV.PHP.S 82 +/- 2% Table 2: Transduction efficiency of AAV.9 versus AAV.PHP.S...figure 2) to allow for efficient IV delivery of virus to the CNS or PNS (figure32, tables 1 & 2). These...that cells are labeled with their own unique color; and 2) low levels of labeling so that a cell’s projections...in tables 1 and 2, with a cell-type specific promoter or enhancer now allows for cell-type-restricted ... overall (GFP+ cells that also stained for a neuronal marker). Check out tables 1 and 2 below for a summary...high dose to maximize the number of cells that are labeled. Part 2: An AAV expressing the inducer (i.e...require the generation of a transgenic mouse. 2. Cell-type-specific restricted transgene expression in...

...pLVX-EF1alpha-SARS-CoV-2-N-2xStrep-IRES-Puro: Expression of the SARS-CoV-2 N protein  pLVX-EF1alpha-SARS-CoV-2-M-2xStrep-IRES-Puro...SARS-CoV-2 M protein pLVX-EF1alpha-SARS-CoV-2-orf3a-2xStrep-IRES-Puro: Expression of the SARS-CoV-2 ORF3a... SARS-CoV-2 protein expression The Krogan lab quickly shared plasmids expressing SARS-CoV-2 proteins at...pLVX-EF1alpha-SARS-CoV-2-E-2xStrep-IRES-Puro) and NSP2 (pLVX-EF1alpha-SARS-CoV-2-nsp2-2xStrep-IRES-Puro...months: Lentiviral preps for expressing SARS-CoV-2 proteins ChR2 for targeted expression DREADDs GFP ...Serotype Name 83899 AAVrg pAAV-mDlx-GCaMP6f-Fishell-2  Additional resources on the Addgene blog Read... results, but also confirm expression in target cells. While the various AAV serotypes can be used to ...

...the image can be acquired more quickly. 2-Photon Microscopy 2-Photon Microscopy attempts to solve two ...Fluorescence microscopy. Nature methods 2, 910-919 (2005). PubMed PMID: 16299476. 2. Conchello JA, Lichtman JW. Optical...provide a 2-10 fold increase in resolution, down to 10s of nanometers. Thick dynamic samples Ex: 3D cell cultures...enhance spatial contrast (i.e. optical sectioning (2) or super-resolution (3). Fluorescence microscopy...photobleaching and a reduction in signal intensity. 2-Photon microscopes restrict excitation (and bleaching...can be seen exiting through the tissue. Therefore, 2-photon allows for deeper imaging into thick tissues...imaging objective that collects fluorescence. Like 2-photon microscopy, only a single focal plane of the...

...Anti-SARS-CoV-2 Nucleocapsid Protein [mBG86], target distinct epitopes on the SARS-CoV-2 nucelocapsid protein...repository of tools for SAR-CoV-2 research. Two new antibodies, Anti-SARS-CoV-2 Nucleocapsid Protein [mBG17...Anti-SARS-CoV-2 Antibodies here! Terry, J.S., et al. (2021). Development of a SARS-CoV-2 nucleocapsid ...domain, PE7 (Figure 2).  PE7 shows enhanced editing efficiencies in a number of cell types at different...Figure 3).      Figure 3: HeLa cells transduced with SARS-CoV-2 nucleocapsid protein (Plasmid #141391...prime editor PE7 Antibodies to facilitate SARS-CoV-2 research New viral vector preps of Voltron2 voltage...editor. Find PE7 plasmids here!     Figure 2: PE7 improves prime editing. A) Schematic of PE7 prime...

...SARS-CoV-2 infects host cells through binding the human angiotensin converting enzyme 2 (ACE2) cell-surface...severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus may acquire mutations to escape ...Figure 1. (A) SARS-CoV-2 RBD DMS Library generation and sequencing. From Starr et al Cell., 2020. (B) Mapping...designed so that the SARS-CoV-2 spike RBD is displayed on the yeast cell’s surface. The Bloom lab used ...Pooled library to study antibody escape in SARS-CoV-2 Recombinant antibodies for aminoacyl tRNA synthetases...viral preps Mapping antibody escape in SARS-CoV-2 by mutational scanning By Seth Kasowitz As multiple... binding ACE2 is the target of most anti-SARS-CoV-2 antibodies. In the library, each plasmid encodes the...

...state-of-the-art methods of sex and gender analysis and 2) provides case studies with concrete examples of how...never considered the sex of their cells. For one thing, cells and cell lines degenerate over time—and sex... effects of using all female cells and tissues, specifically in stem cell therapies. Let’s go back to ...therapeutic capacity of stem cells. Yet, very few researchers consider the sex of their cells when doing experiments—which...sexes in basic research is excellent design), but they used all female stem cells—this was an unconscious... stem cells. Nor did they detect important differences in function between male and female cells. The ...experiment. In the case of stem cells, these factors may include cell type, the disease being treated,...

...interest in the cells you are studying. Read our SPARC blog post for more details. Figure 2: Dense (c... rising intracellular calcium levels within the cell. When levels of Ca2+ are high in the cell, calcineurin...infection (MOI) during cell library construction to ensure that the majority of cells receive one gRNA. The... of a genetically-defined subset of cells. By labeling fewer cells in a population, it’s easier to visualize...lengths, the density of cells that are labeled can be altered to dense (45% of cells labeled), intermediate...or aging cell populations. Figure 3: Micrographs of wild type (top panels) and cells with high...barcodes (iBARs) in their loop region. After infecting cells with sgRNAiBAR library containing lentiviral vectors...

...encyclopedia of Class 2 CRISPR systems with wiki entries describing enzyme activity, experimental considerations...library amplification 1 vector system: lentiCRISPR v2 2 vector system: lentiCas9-Blast and lentiGuide-Puro...and introducing it into cells. Protocol for Genomic Deletions in Mammalian Cell Lines CasPEDIA : An encyclopedia...interest and correspond to edited and non-edited cell populations. For more information, see the bioRxiv...134.7 KB Musunuru CRISPRs in human pluripotent stem cells pCas9_GFP ; gRNA empty vector Link O'Connor-Giles...Bauer Protocol for Genomic Deletions in Mammalian Cell Lines pSpCas9(BB) (pX330) Link Parrott NEBuilder...

...Cre-ERT2 CAG Mammalian Heller 112622 pVHC_PGKneoLox2DTA.2 Venus, Cre-ERT2, targeting vector with MCS for homology...homology arms Mammalian Heller 112623 pEHC_PGKneoLox2DTA.2 Emerald, Cre-ERT2, targeting vector with MCS for homology... arms Mammalian Heller 112624 pmTHC_PGKneoLox2DTA.2 TFP, Cre-ERT2, targeting vector with MCS for homology...arms Mammalian Heller 112625 ptdTHC_PGKneoLox2DTA.2 tdTomato, Cre-ERT2, targeting vector with MCS for ...25;1504(4):467-86. doi:10.1016/0022-2836(81)90375-2. PubMed . Ventura A, Meissner A, Jaenisch R, Jacks...allowing for direct comparison of Cre+ and Cre- cells. While Cre-lox recombination is a very useful tool...promoter like CAG, or expressed only in a subset of cells under a more specific promoter (e.g. Rho-Cre is ...

...Cas enzymes? CasPEDIA is an encyclopedia of Class 2 CRISPR systems with wiki entries describing enzyme...is highly recommended that you determine if your cell type contains the target sequence before using any...

...are largely underutilized compared to their Class 2 counterparts like Cas9. This is attributed in part...bound to DNA, and the break can be repaired by the cell’s endogenous repair mechanisms. Cas3 belongs to the...

...Multiple SARS-CoV-2 variants escape neutralization by vaccine-induced humoral immunity. Cell 184:2372-2383...months, we’ve seen the emergence of several SARS-CoV-2 variants coincide with increased vaccination efforts...They were able to tweak this assay to study SARS-CoV-2 spike variants. “We created a series of spike expressing...relative ability to neutralize.”   Figure 2: Sera used in the study came from individuals who ...This sera was used in the high-throughput SARS-CoV-2 pseudovirus neutralization assay. Neutralization was...viruses are quite genetically distinct from SARS-CoV-2 and its variants. Yet, the B.1.351 variant, which ...effectiveness of current vaccines against SARS-CoV-2 variants, this study was the first to compare so many...

...Illustration of SARS-CoV-2 from Philippa Steinberg for the IGI.   What SARS-CoV-2 plasmids from Ginkgo ...share over 1,850 SARS-CoV-2 plasmids to help academic and industrial labs accelerate research and to help ...request? Currently, plasmids containing all SARS-CoV-2 protein coding genes in E. coli and S. cerevisiae ...bacterial expression systems, Ginkgo synthesized SARS-CoV-2 expression plasmids in a high copy backbone with a...high level expression and purification of SARS-CoV-2 proteins for downstream experiments and offer flexibility...optimized, full length and truncated versions of SARS-CoV-2 S (spike protein) cloned into a CMV driven expression...first deposited set of plasmids containing SARS-CoV-2 protein coding genes in E. coli and S. cerevisiae ...

...comprehensive SARS-CoV-2 virus-human interaction network in which we expressed 26 of the 29 SARS-CoV-2 viral proteins...understanding how SARS-CoV-2 interacts with human proteins. For example, we used HEK293T cells in our first study...all became acutely aware of the threat of SARS-CoV-2. Like many in the science community, we’ve been motivated...identifying these potential human helpers of SARS-CoV-2 (especially the ones that are druggable) is a key ... joining in this worldwide fight against SARS-CoV-2. As we’ve all been affected in various ways by the...details. Our set of constructs comprises all SARS-CoV-2 viral proteins except for Nsp3 and Nsp16, and includes...is going on there!).  Find the Krogan Lab SARS-CoV-2 plasmids (Note: The Nsp6 construct is undergoing QC...

...this year. Designing the donor plasmid Figure 2: Donor plasmid design strategy. A donor plasmid contains...Institute for Cell Science. Learn more by visiting the Allen Cell Explorer at allencell.org and the Allen... hiPS cell lines generated using these plasmids, please visit the Cell Catalog on the Allen Cell Explorer... tagging to elucidate cellular structures. Overexpression methods flood the cell with protein, which can... in human cell lines. Furthermore, the quality control necessary to ensure the edited cells are behaving..., and the stem cell lines we’ve made using them are available as part of the Allen Cell Collection at ...

...chromosomal integration of DNA. ACS Synthetic Biology, 2(9), 537–541. DOI: 10.1021/sb400021j Figure 1: Multiplexing... CRISPR plasmids. Each point represents a single cell, and each color represents the population obtained...

... funding from the Innovative Medicines Initiative 2 Joint Undertaking under grant agreement No 777372....quality of research output, RESOLUTE expects to accelerate the pace of research in the field of SLCs to ...

...change the media (add 2-3 mL of fresh media once a week), and do NOT harvest the cells before at least 10...titer. Amplification (1-2 weeks) - The rAdV-S is used to infect more HEK293 cells and produce more rAdV ...infected cells show this “cytopathic effect.” Amplification can be repeated multiple times (2-4 rounds...creating adenoviral vector constructs. It consists of 2 plasmids: a shuttle vector (in which the transgene...Addgene’s Adenoviral Guide webpage, and references 1, 2.*Pro-Tip*: once the correct recombinant pAdV plasmid...sequenced to confirm its integrity. Initial production (2-3 weeks) - Here you’ll produce the primary recombinant...rounds) at increasing scale over the course of 1-2 weeks. Each round of amplification should result in a 10...

...safely. Recently, many SARS-CoV-2 researchers began pseudotyping the SARS-CoV-2 spike protein into a lower ... et a., 2018).   Figure 2: To pseudotype with the SARS-CoV-2 spike protein, scientists encode...of vesicular stomatitis virus, HIV-2, and human foamy virus in cell lines expressing galactosyl(alpha1...spike protein’s role in infecting cell lines without using intact SARS-CoV-2 virus. Scientists have been using...Particles Produced in Human Cells Are Inactivated by Human Serum. Molecular Therapy 2:218–222 . https://doi....production uses three plasmids: (1) The transfer plasmid, (2) the envelope plasmid, and (3) the packaging plasmid... pseudotyping methods with other viruses in a BSL-2 lab instead of a BSL-3 or BSL-4 lab. For more details...