... for real-time multicolor luminescence imaging.” Proc Natl Acad Sci U S A 112(14) (2015): 4352-6. PMID...improve the brightness of previous Nano-lanterns by 2-4 fold and permit five-color fluorescent imaging! ...activation, you can’t use fluorescence to monitor processes directly affected by light. Long-term light exposure...

...direct reprogramming by transformation-deficient Myc. Proc Natl Acad Sci U S A, 2010. 107(32): p. 14152-7. ...blood are reprogrammed immediately after collection [2]. PBMCs display unique epigenetic signatures that ...PMID: 27882265. PubMed Central PMCID: PMC5118044. 2. Kim, Y., et al., The Generation of Human Induced ... to the process of collecting fibroblasts, which require a more invasive skin biopsy procedure. This becomes...measures, thereby increasing the costs of the procedure. Episomal reprogramming is an ideal and cost-effective...

...neurons with vesicular stomatitis virus vectors. Proc Natl Acad Sci USA 108, 15414–15419. PubMed PMID:...for retrograde transport has been found for AAV1, -2, -5, -7, -8 (Taymans et al., 2007), AAV1 (Hollis et... tropism and axonal transport of AAV serotypes 1, 2, 5, 6, 8 and 9 in the mouse brain. PLoS One 8:e76310...reveal novel hippocampal CA2 circuits. Nat Neurosci 17(2), 269–279. PubMed PMID: 24336151. PubMed Central PMCID... Mechanisms of Motility and Regulation. Neuron 84(2), 292–309. PubMed PMID: 25374356. PubMed Central PMCID...transduction efficiency of recombinant AAV serotypes 1, 2, 5 and 8 in the rat nigrostriatal system. J Neurochem...analysis of adeno-associated viral vector serotypes 1, 2, 5, 7 and 8 in mouse brain. Hum Gene Ther 18, 195–...

...507-17. 10.1016/j.chembiol.2012.02.006. 2 Guntas et al., Proc Natl Acad Sci USA 2015 Jan 6; 112(1):112... the genome of your favorite cells. Ma et al., Proc Natl Acad Sci U S A. 2015 Mar 10;112(10):3002-7....precision in a native context. Chatterjee A et al. Proc Natl Acad Sci U S A. 2013 Jul 16;110(29):11803-8... Zetsche et al., Nat Biotechnol. 2015 Feb 2;33(2):139-42. doi: 10.1038/nbt.3149. Light-inducible ...and on your favourite signalling pathways (Figure 2). They are all available now at Addgene. 1 Lungu...Nihongaki et al., Chemistry and Biology. 2015 Feb 19; 22(2):169-74. High efficiency cloning for enzyme screening...inducible, gene knockdown with shRNA or shRNA-miR30, (2) tamoxifen-inducible CreERT2 gene deletion, (3) simultaneous...

...with the endogenous tRNA-processing system. Xie K, Minkenberg B, Yang Y. Proc Natl Acad Sci U S A. 2015...3570-5. doi: 10.1073/pnas.1420294112. Epub 2015 Mar 2. PubMed. Find the plasmids from this publication ...conserved cellular machinery to process gRNAs. Hijacking tRNA processing to make gRNAs In a recently published...in rice plants. Yang was intrigued by the tRNA processing machinery present in almost every living organism...gRNAs with any 5’ leading nucleotide could be processed, a distinct advantage over other methods that ...is its universality; robust and efficient tRNA processing is common to prokaryotes and eukaryotes. Though...

...immunodeficiency virus type 1 preintegration complexes.” Proc. Natl. Acad. Sci. U.S.A. 89 (1992) 6580–6584. PubMed...PubMed PMID: 8491198. PubMed Central PMCID: PMC413431. 2. Bukrinsky MI, et al. “Active nuclear import of human...

...immunohistochemical reagents. Yamagata M, Sanes JR Proc Natl Acad Sci U S A. 2018 Feb 13. pii: 1722491115...JS, Legube G, Miller KM Genes Dev. 2015 Jan 15;29(2):197-211. DNA Cross-Bridging Shapes a Single Nucleus...would become busy and potentially detract from the process being depicted, we can’t forget that those other...plasmids through Addgene, don’t hesitate to start the process! Addgene depositors who study chromatin The PIs...

...protein expressed by recombinant vaccinia virus. Proc Natl Acad Sci U S A. 1991;88(20):8972-6. Moks T,...take care when using this method.      Fig 2:  A, The polyhistidine tag is a string of histidine...

...Illustration of SARS-CoV-2 from Philippa Steinberg for the IGI.   What SARS-CoV-2 plasmids from Ginkgo ... traditional, QC process. While this has worked well for the early sets of SARS-CoV-2 materials, it will...Bioworks to synthesize and share over 1,850 SARS-CoV-2 plasmids to help academic and industrial labs accelerate...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...

...cancer. These viruses need to be handled in a BSL-2 (or BSL-2+) lab and require additional safety measures.... BSL-1. In BSL-2 labs, eye protection is required any time you have to work with BSL-2 materials, such...respiratory diseases in humans, earning this virus a BSL-2 classification. Lentivirus (a type of retrovirus) ...recommended or required in BSL-1 and required in BSL-2 labs. When working with lentivirus, many organizations...sprayed into the air. For lentivirus and other BSL-2 materials, you’ll need to use aerosol-tight caps, ...percentage of the decontaminate to be used for BSL-2 waste, and of course, if you are working with a substance...plasmid (packaging plasmid). Viral production then proceeds by transfection of all three plasmids into a producer...

...neurons in the adult dentate gyrus and olfactory bulb. Proc Natl Acad Sci U S A. 2013 Mar 19;110(12):E1152-61... 2008 Dec; 18(6): 617–623. Pubmed PMID 19349161.  2. Deshpande A, Bergami M, Ghanem A, Conzelmann KK, ...foundational later on when we studied biological processes, like how our stem cells are housed in specific...G-deleted RABV can divide and replicate (since these processes are mediated by other viral proteins), but the...

...severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus may acquire mutations to escape ... SARS-CoV-2 infects host cells through binding the human angiotensin converting enzyme 2 (ACE2) cell-surface...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...viral spike protein.   Figure 1. (A) SARS-CoV-2 RBD DMS Library generation and sequencing. From Starr... binding ACE2 is the target of most anti-SARS-CoV-2 antibodies. In the library, each plasmid encodes the...and a barcode, and is designed so that the SARS-CoV-2 spike RBD is displayed on the yeast cell’s surface...

...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...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...relying on raster scanning as is done in confocal or 2-photon microscopes. Total Internal Reflection Microscopy...

...comprehensive SARS-CoV-2 virus-human interaction network in which we expressed 26 of the 29 SARS-CoV-2 viral proteins...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 ...just at the beginning of understanding how SARS-CoV-2 interacts with human proteins. For example, we used... 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...

... be extracted from small bacterial cultures (i.e. 2-3 ml) with or without commercially available kits ...ratios measured at 260 and 280 nm should fall ~1.8 - 2 and give scientists an indication of purity versus...contaminants. The 260 and 230 nm ratios should fall ~2 - 2.2, and provide an indication of purity versus ...hydrochloride used during plasmid extraction. Figure 2: Use cloning grade DNA for direct cloning of your ...Transformation grade DNA  Transformation is the process of inserting exogenous DNA in a microorganism. ...contaminants that have been used during the purification procedure. Good indicators of DNA purity are the absorbance...transfection grade DNA. Transfection is the process of inserting an external plasmid in mammalian cells...

...the addition of Cre recombinase (Figure 2). Figure 2: A knockout targeting vector designed to... an idea of the process, we’ll walk through an experiment designed to knock out exon 2 of a given gene...For recombination to occur in a cell, a minimum of 2 kb of sequence homology is required, but 6 to 14 kb...construct to your target cells. After recombination, exon 2 of the targeted gene will be removed from the chromosome...the GFP/NeoR cassette is inserted in place of exon 2. Thus, the targeted gene is disrupted (knocked out...recombination event will remove NeoR only and leave exon 2 floxed, as seen in line 4 of Figure 4. Since the loxP...Gene targeting takes advantage of this natural process to replace a targeted genetic locus with homologous...

... H. J. (2009). Rats! Disease Models & Mechanisms, 2(5–6), 206–210. https://doi.org/10.1242/dmm.002733 ...that has defined our understanding of biological processes and developments. Model organisms often feature...history and characterization. Many proteins and processes naturally found in E. coli have been manipulated.... coli can be used as a model to relate these processes in more pathogenic bacteria (Blount, 2015). Rattus...McCusker and Gardiner, 2011). As many biological processes and signaling pathways that regulate regeneration...humans. Thus, understanding their life cycle and processes with their regenerative abilities is expected ...the most significant and fundamental biological processes. Many of their genetics and developmental patterns...

...determined by both codon bias and folding energy. Proc Natl Acad Sci USA 107:3645–3650 https://doi.org/...Hoogsten base pairs and wobble pairs.  Figure 2: (A) The codon wheel depicts the 64 codons that encodes...

... needing to work with a transfectable cell line. 2) They require larger expression vectors which are ...are also larger than their Cpf1 counterparts. Table 2. Cas9 Multiplexing Options  Multiplexing Method... Cpf1 expression vector.  See the graph in figure 2 for a comparison of editing frequency resulting from...Biotechnology 35.1 (2016): 31-34. Pubmed PMID: 27918548. 2. Zetsche, B., Gootenberg, J., Abudayyeh, O., Slaymaker...Cpf1 Is a Single RNA-Guided Endonuclease of a Class 2 CRISPR-Cas System. Cell,163(3), 759-771. PubMed PMID...like Csy4, or the inclusion of processing signals, such at tRNAs, to process the array into functional crRNAs...Cpf1’s ability to process its own pre-crRNA arrays simplifies the crRNA cloning process. For cloning, Zetsche...

...Biol. 2015. PubMed PMID: 25895059 Barnea, et al. Proc Natl Acad Sci U S A. 2008. PubMed PMID: 18165312...Science. 2013. PubMed PMID: 24136966 Nyerges, et al. Proc Natl Acad Sci USA. 2016. PubMed PMID: 26884157 ...generating protein chains or clusters. Veggiani, et al. Proc Natl Acad Sci U S A. 2016. PubMed PMID: 26787909...backbone, which expresses the gRNA from a Drosophila U6:2 promoter and Cas9 from the actin 5C promoter. Addgene...backbones for lentivirus LV 1-5 and retrovirus RV 2-5 were constructed. The authors then used GMAP assembly...engineering (MAGE) (1). MAGE uses recombineering (2) to simultaneously incorporate multiple single-strand...number of tandemly expressed gRNAs respectively) and 2) the size of the deletion. Smaller deletions generally...