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Showing: 1 - 13 of 13 results
  1. Fluorescent Proteins 101: GFP Fusion Proteins - Making the Right Connection

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    Blog Post
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    Dec. 23, 2019, 2:33 a.m.
    ...experience, the best site for the insertion is a loop in between secondary structure (beta-sheets or alpha...successful insertion of a fluorescent protein are the tagging of Ccd42 (Bendezú et al., 2015) and the alpha...GFP and its homologues have a beta-barrel structure....One of the limitations is the size of GFP: ~240 amino acids or about 28 kDa....Despite these potential issues, GFP has been used successfully in countless fusion proteins....For instance, GFP may occlude catalytic sites or obstruct binding sites or motifs that are necessary...Keep the linker sequence short In the early days of GFP applications, many were concerned with steric...In fact, for several FRET biosensors (yellow cameleon 3.60, EPAC, and Galphai), the C-terminus and N-terminus...
  2. Plasmids 101: SunTag and Fluorescent Imaging

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    Blog Post
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    Oct. 20, 2019, 9:08 p.m.
    ...To increase expression, Tanenbaum et al. modified the GCN4 sequence to increase its alpha-helical structure...gfp...Initially, Tanenbaum et al. observed some GFP aggregation, which they reduced by using superfolder GFP...the effects of mitoNEET-SunTag-GFP....has a molecular weight of 1,400 kDa vs 24 kDa for GFP alone....In SunTag nomenclature throughout the rest of this blog post, GFP refers to sfGFP-GB1.  ...GCN4 recruits GFP fused to the cognate scFV antibody, which is expressed from a separate plasmid....In SunTag nomenclature throughout the rest of this blog post, GFP refers to sfGFP-GB1.  ...(sfGFP) with the small solubility tag GB1....
  3. Plasmids for Endogenous Gene Tagging in Human Cells

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    Blog Post
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    Oct. 20, 2019, 8:53 p.m.
    ...87420 PXN-EGFP AICSDP-5 EGFP Paxillin Matrix Adhesions 87421 TUBA1B-mEGFP AICSDP-12 mEGFP Alpha-tubulin...ve created and tested plasmids that use CRISPR/Cas9 to endogenously tag a wide variety of genes with GFP...For a C-terminal tag, as shown in the example above, we inserted a linker and GFP tag at the end of the...Plasmids, plasmids everywhere To date, we have created 10 plasmids that can be used to introduce a GFP...The whole segment—a GFP tag and 1kb of homologous DNA flanking both sides (about 2.7kb total)—is inserted...This results in the insertion of the GFP tag precisely where we need it, in this example, at the C-terminus...Matrix Adhesions 87421 TUBA1B-mEGFP AICSDP-12 mEGFP Alpha-tubulin Microtubules 87422 LMNB1-mEGFP...87424 DSP-mEGFP AICSDP-17 mEGFP Desmoplakin Desmosomes 87425 ACTB-mEGFP AICSDP-16 mEGFP Beta-actin...Actin 87426 SEC61B-mEGFP AICSDP-10 mEGFP Sec61-beta Endoplasmic Reticulum 87427 FBL-mEGFP...AICSDP-14 mEGFP Fibrillarin Nucleolus Coming Soon MYH10-mEGFP AICSDP-24 mEGFP Non-muscle myosin...AICSDP-13 mEGFP Nuclear laminB1 Nuclear envelope 87423 TOMM20-mEGFP AICSDP-11 mEGFP TOM20 Mitochondria...Addgene ID Plasmid Allen Institute ID Tag Protein Structure 87420 PXN-EGFP AICSDP-5 EGFP Paxillin...heavy chain IIB Actomyosin bundles 87429  TJP1-mEGFP AICSDP-23 mEGFP Tight junction protein ZO1 Tight...
  4. Fluorescent Proteins 101: When GFP lets you down

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    Blog Post
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    Dec. 14, 2019, 3:11 a.m.
    ...In this way, we (and others) have successfully generated several functional heterotrimeric G-alpha subunit...There are numerous GFP variants. Here I use the name ‘GFP’, which refers to mEGFP....GFP is big GFP is a 28 kDa protein that resembles a cylinder with a length of 4.2 nm and a diameter of...GFP can only be used for tagging proteins GFP fusion proteins are made by connecting its DNA code to...GFP is sensitive to acid The chromophore of GFP can exist in a protonated and a deprotonated state (Tsien...GFP is not tolerated in my fusion It is standard practice to attach the GFP to the N- or C-terminus of...GFP fluorescence overlaps with autofluorescence The cyan light that is used to excite GFP, may also excite...There are numerous GFP variants. Here I use the name ‘GFP’, which refers to mEGFP....to create enhanced GFP (EGFP), a variant with improved brightness (Tsien, 1998)....This variant was made by introducing two mutations (F64L, S65T) in the original jellyfish GFP (AvGFP)...Another mutation (A206K) is necessary to generate the strictly monomeric EGFP variant, mEGFP (Zacharias...
  5. Plasmids 101: The Promoter Region – Let's Go!

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    Blog Post
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    Dec. 2, 2019, 1:42 p.m.
    ...EF1a General expression mRNA Strong mammalian expression from human elongation factor 1 alpha Constitutive...Browse All Plasmids 101 Posts Learn about Mammalian Vectors Read about Reporter Gene like Luciferase and GFP...
  6. New and Upcoming Viral Vectors - May 2020

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    Blog Post
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    May 28, 2020, 1:15 p.m.
    ...The GFP dependent FLPo, Flp-DOG (75469), is also available in AAV1....few months: Lentiviral preps for expressing SARS-CoV-2 proteins ChR2 for targeted expression DREADDs GFP...now expanded to offer many of our control plasmids in new serotypes, as well as a new Cre dependent GFP...CRE-DOG constructs 69570 and 69571 together encode the N and C terminal segments respectively of a GFP...system takes advantage of the extensive supply of transgenic reporter lines that supply a variety of GFP...expression patterns, thus allowing you to further restrict gene activation to cells expressing both Cre and GFP...4 Controls Plasmid Serotype Name 114471 AAV2 pAAV-Ef1a-fDIO mCherry 83900 AAVrg pAAV-mDlx-GFP-Fishell...preps for expressing the E protein (pLVX-EF1alpha-SARS-CoV-2-E-2xStrep-IRES-Puro) and NSP2 (pLVX-EF1alpha-SARS-CoV...of the SARS-CoV-2 N protein  pLVX-EF1alpha-SARS-CoV-2-M-2xStrep-IRES-Puro: Expression of the SARS-CoV...The following lentiviral preps are now available: pLVX-EF1alpha-SARS-CoV-2-N-2xStrep-IRES-Puro: Expression...-2 M protein pLVX-EF1alpha-SARS-CoV-2-orf3a-2xStrep-IRES-Puro: Expression of the SARS-CoV-2 ORF3a Lentiviral...50457 AAV1 pAAV-hSyn-DIO-EGFP Bryan Roth 114472 AAVrg pAAV-hSyn-mCherry Karl Deisseroth 83895...
  7. 28 Hot Plasmid Technologies from 2015

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    Nov. 20, 2019, 2:36 p.m.
    ...(tubular ER) Microtubules mCherry-alpha-tubulin Early Endosome Vacuolar Compartment mCherry-Rab5...(GTPase in division) GFP-Mff (outer membrane protein) Late Endosome mCherry-Rab7A GFP-Rab7A  ...BFP-Rab5 GFP-Rab5B Vacuolar & Budding Compartment GFP-Rab4B Mitochondria mito-BFP mCherry-Drp1...protein that binds to the coding region of a reporter mRNA via a PP7 coat protein (NLS-PCP-GFP) a RFP...By fusing these dCas9s with one of three fluorescent proteins (GFP, BFP and mCherry), the authors were...During the first round of translation, the GFP proteins that are bound to the coding region are displaced...Broccoli is an RNA aptamer that acts as a GFP mimetic and induces fluorescence in the presence of DHFBI...Co-expression of LOC8R-PSmOrange and LOC8R-PAGFP provides a novel strategy for assaying the dynamics...to the protein of interest are recognized and thus fluorescently labeled by scFv antibodies fused to sfGFP...
  8. CRISPR 101: Epigenetics and Editing the Epigenome

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    June 25, 2020, 11:45 a.m.
    ...Transcriptional repression DNA Methyltransferase 3 Alpha (DNMT3A) Vlatka Zoldoš’ lab has deposited pdCas9...Co-expression markers EGFP and PuroR enable sorting and selection of transduced cells....-DNMT3A-EGFP and pdCas9-DNMT3A-PuroR for targeted cytosine methylation in mammalian cells....DNA Methyltransferase MQ1 Margaret Goodell's lab has deposited pcDNA3.1-dCas9-MQ1(Q147L)-EGFP, a fusion...over a period of several days, as seen with other epigenome-editing tools. pLV hUbC-dCas9-MQ1(Q147L)-EGFP...
  9. Plasmid Tools for Microbiome Studies

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    Jan. 6, 2020, 4:12 p.m.
    ...also contain the RP4 transfer origin as well as a selectable marker and a desired genetic payload (ex: GFP...from the microbial community) reveals the RBS and allows translation of the reporter gene resulting in GFP...The plasmids could also be transformed into Alphaproteobacteira, Betaproteobacteria, and Gammaproteobacteria...
Showing: 1 - 13 of 13 results