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We narrowed to 956 results for: REP

Showing: 301 - 320 of 956 results

  1. Components of CRISPR/Cas9

    Type
    Blog Post
    ... (NM) 3' NNNNGATT  Streptococcus thermophilus (ST) 3' NNAGAAW  Treponema denticola (TD) 3' NAAAAC...their functions and simply called the repeated elements “direct repeats” and the unique stretches of DNA between... Species/Variant of Cas9 PAM Sequence Streptococcus pyogenes (SP); SpCas9 3' NGG SpCas9 D1135E... CRISPR arrays in bacterial genomes consist of repeated elements separated by unique sequences. When researchers...years of research, we now know that each direct repeat, combined with its adjacent spacer, ultimately ...ultimately encodes a single crRNA. The direct repeat regions contain sequences required for processing pre-crRNA...each individual crRNA. Bottom line, the “direct repeat region” combined with the tracrRNA forms the scaffold...

  2. Plasmids 101: Mammalian Vectors

    Type
    Blog Post
    ... a bacterial origin of replication (ORI) will not allow for plasmid replication in mammalian cells, for... their bacterial counterparts, including how replication occurs and whether selection is necessary for...not used for general plasmid selection. Instead, reporter genes are often employed to easily monitor transfection...expression levels in the cells. Ideally, the chosen reporter is unique to the cell, is expressed from the plasmid...assess transfection success. GFP is often used as a reporter and we will be covering this ever-versatile fluorophore...the nuclear genome, in the absence of plasmid replication, the foreign DNA will be degraded or diluted ...always mission critical to make sure the plasmid replicates in the cell, as many of these experiments are...

  3. Multiple Plasmids at a Low Price: Kits from Addgene

    Type
    Blog Post
    ...designed repeat protein combinatorial library by in vivo functional screen." Scientific reports 5 (2015...fusions reported in the literature include RNA detection, RNA cleavage, and translational repression or activation...of the many useful kits you can order from the repository: FX Cloning Kit BIOFAB Plasmid Set pCri System...including antibiotic efflux pumps and ankyrin seven-repeat proteins: Sjuts, Hanno, et al. "Molecular basis...strengths centered within GFP/RFP bicistronic reporters characterized in Cambray et al., 2013. These terminators... an Orthogonal Ribosome-Controlled Fluorescent Reporter."ACS chemical biology 11.1 (2015): 31-37. pCri...expression systems; the Cell-free expression test kit represents a set of 88 clones in a pCellFree vector that...

  4. The Addgene Antibody Data Hub

    Type
    Blog Post
    ...Antibody Data Hub. Data reports  Each antibody submission is used to create a data report, which contains experimental...that information in the report as well.    Figure 3: An antibody data report, showing the results of ...methods section for the data report shown in Figure 3.   At the end of the report, you’ll find a results section... the data report shown in Figure 3. The antibody is rated "Pass". Curating the data reports Just like... page of the Addgene Data Hub.   Antibody data reports can be searched and sorted by gene/target, name...information is then formatted and published as a report with a DOI for citation. If you’re considering ...

  5. CRISPR 101: Epigenetics and Editing the Epigenome

    Type
    Blog Post
    ...epigenome may make certain processes, like cellular reprogramming of iPSCs, more efficient. In general, epigenetic...However, several proteins in the DNA oxidation and repair pathways work downstream of Tet1 to restore the...of Tet1 alone, or coupled with DNA oxidation and repair factors that allow for increased gene activation... other Tet1 delivery systems. Transcriptional repression DNA Methyltransferase 3 Alpha (DNMT3A) Vlatka...methylation is associated with transcriptional repression. Created with BioRender.com. DNA Methyltransferase...as a kit from Addgene.   Find Ready-to-Use Viral Preps for Epigenetic Modification Why use epigenetic modifiers...activates transcription, whereas dCas9-KRAB fusions repress transcription. Both of these methods also recruit...

  6. Hot Plasmids - October 2020

    Type
    Blog Post
    ...a subset of the new plasmids and viral preps in the repository through our hot plasmids articles. These...proliferate. Eastman et al. Cell Reports 2020. https://doi.org/10.1016/j.celrep.2020.107804 The CRISPR corner...to provide ready-to-use viral preps. Here are some of the new viral preps from recent months: The Dolcetto...Addgene Sc++ is a codon-optimized, engineered Streptococcus canis Cas9 nuclease that has a broad 5′-NNG-...inhibition pooled library is now available as lentiviral preps in addition to the pooled library format! The library...inhibits over 18,000 human genes. Find the lentiviral preps at Addgene hSyn1-SIO-stGtACR1-FusionRed is a Cre-dependent...channelrhodopsin. It’s now available as AAV5. Find the AAV prep here The red-shifted calcium sensor jRGECO1a is...

  7. Plasmids 101: FLEx Vectors

    Type
    Blog Post
    ...invert the reporter and GeneR fragment. Then the second recombination event would excise the reporter from ...Spatio-temporal expression of optogenetic reporters: Since most optogenetic reporters are delivered using AAV injection...transduced reporter gene. Generating a FLEx switch to control expression of an optogenetic reporter would ...Cre-dependent reporter expression, the potential for background expression of the reporter will be greatly...shown in Figure 2, the cassette should contain a reporter DNA sequence (such as GFP) in the sense orientation...the two pairs of target sites. By flanking the reporter with one loxP site and one lox511 site in the ...would ensure that the reporter remains silent until a cell or tissue-specific Cre is provided. Since the FLEx...

  8. Illuminating Epigenetics with A FRET Based Biosensor

    Type
    Blog Post
    ... versions of this reporter to target the K9 (pcDNA3-K9 histone methylation reporter) and K27 (pcDNA3-K27...methylation reporter) positions on histone H3, which are known to be involved in repression and X-inactivation...The K9 reporter uses the HP1 chromodomain to recognise lysine 9 methylation, while the K27 reporter uses... promoter region of tumor suppressor genes can repress their transcription and inactivate them. Global... methylation in living cells. Essentially this reporter construct consists of one substrate (the histone-derived...methylation of these sites was controlled, the reporter consistently showed significantly different FRET...states. Thus, this system is a reliable tool in reporting methylation status of these sites. This new technology...

  9. CRISPR 101: Mammalian Expression Systems and Delivery Methods

    Type
    Blog Post
    ...Traffic Light Reporter System: Can evaluate lentiviral component delivery as well as genome repair by non-homologous...in genome target sequence into EGFP reporter. Target DNA reporter system: Can evaluate component delivery...or knock-in a gene? Do you want to activate or repress gene expression? Are you trying to create a single...mutation? Do you want to create a fusion with a reporter protein such as GFP? All of these outcomes can... can be used in conjunction with activator or repressor domains to control gene expression. Base editors...detecting insertions/deletions using a mismatch repair assay? Or is PCR followed by gel electrophoresis...application. If you want to include a homology directed repair (HDR) template to create knock-ins, point mutations...

  10. Technical Design of a Western Blot

    Type
    Blog Post
    ...primary antibody required. Reporters There are two main classes of reporters: chemiluminescence enzymes...are more consistent lot-to-lot. The highest reproducibility comes from recombinant antibodies (rAbs), which...that target the same protein — can be used to reproduce the benefits of polyclonal antibodies. Because...you need a specific conjugate, and changing your reporter molecule requires either re-conjugating or re-...-purchasing your primary antibody with the new reporter. Finally, each new conjugate will have to be tested...binding to your protein of interest. Protein preparation Before you can blot your proteins, you’ll need... is a sturdy membrane that can be stripped and reprobed multiple times. It doesn’t degrade during long-term...

  11. DNA Quantification

    Type
    Protocol
    ...NanoDrop to measure your samples, place 1-2µL of mini-prepped DNA onto the pedestal. Close the lid and click ...280 column (A good purity ranges from 1.80-2.00). Repeat for each sample. Notes: Keep in mind that despite...

  12. Build Your CRISPR Vocabulary

    Type
    Blog Post
    ...RNA” base pairs to the repeat section of the crRNA, forming the repeat:antirepeat duplex. tracrRNAs also...Clustered Regularly Interspaced Short Palindromic Repeats. That contrast of simplicity and complexity is ...array. The CRISPR array is composed of identical repeat sequences and variable spacer sequences. The array...transcribed and processed into crRNAs, each including one repeat and one spacer. In bacteria, these crRNAs are bound... bound by Cas proteins (Cas9 shown here). The repeat sequence base pairs with the tracrRNA, and the spacer... of these arrays, with alternating spacers and repeats, was the original impetus for studying CRISPR. ...array, these sequences are called protospacers. Repeats: Bacterial DNA that appears between each spacer...

  13. Plasmids 101: Gene, enhancer, and promoter trapping

    Type
    Blog Post
    ...fundamentally the reporter construct monitors the expression of a nearby gene. The reporter gene serves as... screens to introduce reporters throughout an organism’s genome. These reporters have historically been...stimulate the expression of the reporter. The system is made up of a reporter gene flanked by a minimal promoter...promoter – all rely on the random integration of a reporter construct in close proximity to an endogenous ...example, if green fluorescent protein (GFP) was the reporter gene and green cells were observed in the kidneys... for with an uncharacterized gene. The actual reporter construct can be something phenotypic (such as...promoter. The system is composed of a promoter-less reporter which needs to integrate within an exon of a gene...

  14. Twenty Years of Sharing: Expanding Accessibility

    Type
    Blog Post
    ... materials in our repository. We started to increase accessibility to our repository by making it easier...Addgene opened in 2004, we focused on creating a repository that made it easy to share plasmids, both for...to — so far, we’ve shipped materials from our repository to an amazing 111 countries (Figure 1)! But while...that temperature-sensitive materials like viral preps and libraries were kept at the appropriate temperature...Systems AAV Plasmids Blogs Origin of Replication What is a Plasmid? How to Design Your gRNA for...Addgene blog in 2023. Views from a country are represented in blue; darker blue indicates more views from...YouTube channel, 2023. Views from a country are represented in blue; darker blue indicates more views from...

  15. Hot Plasmids: Fall 2024

    Type
    Blog Post
    ...the new plasmids, antibodies, and viral preps in the repository through our Hot Plasmids articles.  Here's...gene disruption in M. abscessus  Bioluminescent reporter of mechanical forces   Epitope tag antibody ...Western Blot for Anti-V5 [IPI-SV5-Pk1]. Addgene Report, https://doi.org/10.57733/addgene.l0ulu4. Hanke...through eight stages of embryonic development. Representative images of single focal planes through the middle...across species due to differences in natural DNA repair pathways and spontaneous mutations.  Here, the ...results in either bacterial death or indels after DNA repair. B) M. abscessus is first transformed with Cas9...an integrative plasmid containing tetracycline repressor TetR and resistance selection marker KanR. This...

  16. Plasmids 101: The Promoter Region – Let's Go!

    Type
    Blog Post
    ...resistance gene and the origin of replication are necessary to replicate plasmids and make sure the cells... operon Inducible by arabinose and repressed catabolite repression in the presence of glucose or by competitive...transcribed promoters Inducible with galactose; repressible with glucose  Can be used independently or together...mRNA Yeast promoter for alcohol dehydrogenase I Repressed by ethanol Full length version is strong with ...expression Promoter from E. coli tryptophan operon Repressible Gets turned off with high levels of cellular ...lac operon Constitutive in the absense of lac repressor (lacI or lacIq). Can be induced by IPTG or lactose...expression. lacIq mutation increases expression of the repressor 10x, thus tightening regulation of lac promoter...

  17. March for Science

    Type
    Blog Post
    ...Democrats more often than Republicans, the March is not about Democrats or Republicans (16). This is a Pro-...contact your representatives in case you would like to talk with them about science: Representatives: To find...contact information for your representative in the House of Representatives all you need to know is your...in this article are those of the author do not represent a formal stance taken by Addgene or its staff....Climate change Vaccines GMOs Transgender issues Reproductive health Immigration Pollution Healthcare Science...Legislatures: To find the website to contact representatives at the state level you can click on your state...

  18. Addgene: The First Twenty Years

    Type
    Blog Post
    ...ready-to-use viral vector preps. Today, that number has grown to over 114,600 viral preps. And in 2025, we’ll...service would allow people to add a gene to a repository others could request from — Addgene!  Addgene...meant we could grow accessibility alongside our repository.  “I’m sure you hear it all the time, but Addgene...took almost six years to reach our first major repository milestone — 10,000 plasmids deposited and available...conferences. And when CRISPR arrived in 2012, our repository was rapidly able to support sharing of CRISPR...milestone! We were thrilled to see the impact our repository was having on the scientific community, but it...-quality service, open data, and increased reproducibility of materials. In March 2022, we launched our...

  19. Hot Plasmids April 2018 - Protein Degradation, Nanoscopy, FIRE-Cas9, and Yeast Expression Tools

    Type
    Blog Post
    ...random variation, or noise, in reporter expression. To do so, one places reporter expression under the control...expression of a reporter protein, it is possible to change the amount of noise in reporter gene expression...highlight a subset of the new plasmids in the repository through our hot plasmids articles. These articles...deliver the target antibody. TRIM-Away reagents and reporters are available from Addgene - we hope they will... been fused to a variety of protein modules to repress or activate gene expression, sometimes incorporating...lab has deposited plasmids encoding fluorescent reporters and other proteins controlled by GAL4 and ZIF268...progesterone controlled transcription factor as well as reporters specific to each transcription factor. They then...

  20. Working with Nuclear Receptors

    Type
    Blog Post
    ...and utilization - and on self-propagation by reproduction. Both physiological processes are controlled...ligands. Nuclear receptors and their ligands NRs represent the largest family of transcription factors found...expression of genes involved in a broad range of reproductive, developmental, metabolic, and immune response...more NRs for efficient and timely activation or repression of gene expression (see www.nursa.org for a curated...Coregulators can function as coactivators and corepressors depending upon a context defined by ligand, ... third phase of funding, the NURSA Consortium represents a unique funding model in nuclear receptor research...the development of a vast number expression and reporter plasmids and the free exchange of these vectors...
Showing: 301 - 320 of 956 results