... Plasmid Collections Mammalian RNAi Mammalian shRNA Tools for RNAi Browse Plasmids Resources...Plasmids for mammalian RNA interference (RNAi) experiments at Addgene; browse by constitutive lentiviral...mutations in therapeutic settings. As a tool in mammalian cell systems, silencing is achieved through the...., & Feng, L. (2023). A comprehensive review of small interfering RNAs (siRNAs): mechanism, therapeutic...

...synthetic biology plasmids for use in mammalian systems. Mammalian Plasmids Search the table by keyword...plasmids for use in mammalian systems. Plasmid... Plasmid Collections Synthetic Biology Mammalian Synthetic Biology...Biology: Mammalian SynBio Resources SynBio Home Browse All SynBio Assembly Standards SynBio Blog Genome Engineering...

... CRISPR Mammalian Expression CRISPR Plasmids: Mammalian Expression The... that function in mammalian cells can be used to attentuate RNA levels. In mammalian systems, Cas13a does...or cloning method. To isolate mammalian vectors, simply type "Mammalian" into the search bar. When using...designed for use in mammalian systems. CRISPR...CRISPR plasmids have been designed for use in mammalian expression systems. Cut Fully functional CRISPR...Libraries gRNAs Empty gRNA Vectors gRNAs By Species Mammalian Bacteria Drosophila Plant C. elegans Yeast Zebrafish... C->T (or G->A on the opposite strand) within a small editing window specified by the gRNA. Adenine base...

...Genomic Deletions in Mammalian Cell Lines CRISPR: Protocol for Genomic Deletions in Mammalian Cell Lines Other...by a small indel at the sgRNA target site. Design additional primers to analyze for scarring (small indels...Experiments (JoVE) about genomic deletions in mammalian cell lines using CRISPR/Cas9 CRISPR...of CRISPR/Cas9 to create genomic deletions in mammalian cell lines. Along with the video, you can find...Article Citation Generation of Genomic Deletions in Mammalian Cell Lines via CRISPR/Cas9. Bauer DE, Canver MC...potential advantages as compared to single-site small indels given the efficiency of biallelic modification...loss-of-function studies of genes and genetic elements in mammalian cell lines. Protocol For referenced Tables and...

...compartment (chloroplastic, cytoplasmic, glyoxysomal, mitochondrial, peroxisomal) Mutation (specific mutation, "...Browse a diverse set of wild-type malate dehydrogenase genes and watermelon mutant clones for your course-based... Depositor Collections MDH CUREs Community Malate Dehydrogenase CUREs Community (MCC) Collection ...Experiences (CUREs) (Link opens in a new window) The Malate Dehydrogenase CUREs Community (MCC) (Link opens...Experiences (CUREs) centered on a single enzyme, malate dehydrogenase (MDH). The MCC is a national group...Visualization to Develop Student Hypotheses in a Malate Dehydrogenase Oriented CURE. Callahan KP et al....learning and attitudinal outcomes: A study of the Malate dehydrogenase CUREs Community (MCC). DeChenne-Peters...

...synthetic vectors and cultured mammalian cells. Of course, these mammalian vectors must be compatible with...discernable effect on mammalian cells. In this blog post we will discuss how mammalian plasmids differ from...supplement in mammalian cell culture to suppress bacterial growth, and is not appropriate for mammalian selection...Although plasmids do not naturally exist in mammals, scientists can still reap the benefits of plasmid-based...ORI) will not allow for plasmid replication in mammalian cells, for example, and a toxin that kills bacteria...for transfected cells. Before getting into the mammalian plasmid components, it may be useful to describe...introducing genetic material (such as plasmids) into mammalian cells, a process called transfection. Transfection...

...yellow) - 8.0 (red)). Blood pH normally lies around 7.35-7.4 so mammalian cells are sensitive to pH. Anything...Cell Culture Cloning Mammalian Cells with the Agarose Method Plasmids 101: Mammalian Vectors Resources ....org Find mammalian plasmids for your research Find viral vectors for infecting mammalian cells ...flowing in. The vent for the in-flowing air is normally visible and you should always ensure nothing is...cross-contamination can be identified. Even if some mammalian cells lines have similar morphologies, being familiar...

...because of a small, avoidable mistake. Or worse, when you're trapped in a pattern of small errors that ... when you’re in the lab and you're sweating the small stuff? Identifying the Issue You can’t fix an experiment...your prep an investment in accuracy.  Bench setup Small adjustments in the physical organization of your...reduce errors as you work, and the possibilities for small, helpful changes are endless.    Fig. 2... Just like any other experimental approach, use small, incremental changes to understand what works and...

...each clone in case of contamination. We do not normally use antibiotics. Lastly, and perhaps obviously... The Challenges of Cell Culture Plasmids 101: Mammalian Vectors Resources on Addgene.org Check Out Our...Plasmids Find Mammalian Expression Vectors ... mammalian cells... mammalian cell clones...

...can help you make an informed decision. Mammalian cells Mammalian cells are a great option for viral production...you. Table 1: Mammalian versus insect cells for viral production Consideration Mammalian cells Insect...multiple different cell types, including plants, mammals, and insects. In this post, we will compare two...two of the most widely used systems, mammalian and insect cells. Whether you’re new to in-house virus production...production, particularly for labs that already do mammalian cell culture. HEK-293T cells are the most common... AAV, and adenoviruses can all be produced in mammalian cells. However, it is much more common to make...capacity of 8–10 kb. Time Viral production in mammalian cells is relatively quick. After transfection,...

...Born to a family in Malawi that pushed science education, Kingdom Kwapata grew up to be one of the best...among 100,000 eligible to attend the University of Malawi where he focused on agricultural engineering through... Kingdom on opportunities to go into science in Malawi 4:36-5:46 : How Kingdom ended up working in the...University 9:36-10:29 : Kingdom's transition back to Malawi and Lilongwe University of Agriculture and Natural...difficulties surrouding and need for biotech research in Malawi 11:41-14:53 : Kingdom's current projects producing...projects - HIV inhibitors, biofuels, challenges in Malawi, and more 23:45-28:07 : How Seeding Labs is helping... malawi...

... also confines channelrhodopsin activation to a small volume of brain tissue (~1 mm3). Current channelrhodopsins... To make channelrhodopsin-based optogenetics minimally-invasive, Frances Arnold’s lab and Viviana Gradinaru...channelrhodopsins paired with a systemic AAV allow for minimally-invasive optogenetics While ChRgers performed ...in mice to see if they were suited for in vivo minimally invasive optogenetics. Both experiments delivered...paired with PHP.eB’s systemic delivery enables minimally-invasive gene delivery and optogenetic excitation...learning-guided channelrhodopsin engineering enables minimally invasive optogenetics. Nat Methods 16:1176–1184...

... design and optimize. Today we’ll cover the normalization and loading controls needed for relative quantification...In relative quantification, the POI bands are normalized to the loading control: the intensity of each...resulting fraction is reported. Figure 2: Normalization with loading controls. Under each lane, the ...the loading controls are equally strong, so the normalized value simply reports which POI band is stronger...is weaker in condition 1 than condition 2, the normalized values for the two conditions are equal. Created...Created with BioRender.com. This kind of normalization is a nearly ubiquitous tool, but proceed with...your western blot, and depending too much on normalization can amplify other errors in experimental design...

...replicated many of the properties once exclusive to small-molecules: red-shifted spectra, ion sensitivity,...improve the photon budget. Of course, “reverting” to small molecule dyes seems daunting—no one wants to spend...that reacts specifically and irreversibly with a small substrate ligand motif attached to a fluorophore...These modified antibody fragments bind and enhance small molecule fluorogens (Figure 1e). Stains – These..., no genetically encoded tag is required as the small-molecule binding motif targets endogenous protein...the system. A particularly desirable property for small-molecule labeling strategies is fluorogenicity, ...flurry of activity and the majority of the classic small molecule fluorophores, such as rhodamines (Figure...

...post we’ll provide an overview of some CRISPR mammalian expression systems, the typical applications for...what CRISPR components you will need to deliver. Minimally, one or more sgRNAs and Cas9 are required for ...those CRISPR components into your target cells.  Mammalian CRISPR expression systems Each model system has...Expression System Components of System Application Mammalian expression vector Promoter driving Cas9 expression...or stable expression of Cas9 and/or gRNA in a mammalian cell line that can be transfected at high efficiency...expression of Cas9 and/or gRNA in a wide variety of mammalian cell lines. Useful for difficult to transfect ...degraded within the cell.   Delivery methods for mammalian cell lines As mentioned above, the expression ...

... them 17-26% smaller than other Cas13s and a whopping 33% smaller than Cas9! Their small size makes then...specific editor in mammalian cells Not all Cas13d proteins are functional in mammalian cells, but Konermann... easy to package in low-capacity vectors! These small, but mighty type VI-D enzymes are the latest tools... does not occur when Cas13d is expressed in a mammalian system. Since Cas13d is functionally similar to...compared CasRx to multiple RNA regulating methods: small hairpin RNA interference, dCas9-mediated transcriptional...

...CRISPR article "Generation of Genomic Deletions in Mammalian Cell Lines via CRISPR/Cas9" from the Journal of...of CRISPR/Cas9 to create genomic deletions in mammalian cell lines. Below Bauer and Canver discuss the...genomic deletions We were inspired to produce intrachromosomal deletions based on the experiments of Kim ...References: 1. Lee HJ, Kim E, Kim JS. Targeted chromosomal deletions in human cells using zinc finger nucleases...palindromic repeats (CRISPR)/Cas9 nuclease system in mammalian cells. J.Biol.Chem. 2014 Aug 1;289(31):21312-21324... mammalian cell...

...to the XFP in the construct, dBrainbow includes small epitopes fused to each XFP. Secondary antibodies...signal. Imaging endogenous fluorescence in live animals only permits visualization of two colors. Similarly...protein microinjection or crossing of Zebrabow animals to an inducible Cre line. The Schier group developed...cell populations using the Confetti mouse For mammalian systems, the Clevers lab developed the Confetti...expression in the Ubow mouse Another murine reporter animal, the Ubow mouse (Ghigo et al., 2013), was developed...RFP are not present at the same frequencies. The small population size of CFP++ cells permitted researchers...

...virus- a non-integrating RNA virus Episomal vectors- extrachromosomally maintained DNA plasmids that express...Reprogramming efficiency of PBMCs with episomal vectors Episomal vectors are based on the Epstein-Barr ... pCXLE episomal vector system generated by the group is an optimized mixture of three episomal plasmids...expression from the other episomal plasmids. When used on PBMCs, the pCXLE episomal vector system increases...reprogramming purposes. In this post, I’ll describe an episomal vector system that increases reprogramming efficiency...thereby increasing the costs of the procedure. Episomal reprogramming is an ideal and cost-effective method...encoding reprogramming factors behave as separate extrachromosomal elements in the nucleus of the target cell...

... to try to determine whether simpler and maybe smaller Cas systems could exist in nature. To do this,... Institute for uncharacterized genes that were proximal to a CRISPR array and the universal CRISPR integrase...cas4, and a new gene cas14. cas14 encodes for a small Cas protein (40-70 kDa), which is half the size ...

...pcDNA3-EGFP - Mammalian Expression pCAG-GFP - Mammalian Expression pLV-eGFP - Mammalian Lentiviral Expression...A206K) pmEGFP-1 - Mammalian Expression mEGFP-N1 - Mammalian Expression mEGFP-C1 - Mammalian Expression mEGFP-pBAD...pCAG-DsRed2 - Mammalian Expression DsRed2-N1 - Mammalian Expression DsRed2-C1 - Mammalian Expression DsRed2...pCAG-HcRed - Mammalian Expression HcRed1-N1 - Mammalian Expression HcRed1-C1 - Mammalian Expression dKatushka...pQC mPlum IX - Mammalian Expression mPlum-N1 - Mammalian Expression mPlum-C1 - Mammalian Expression mPlum-pBAD... Sirius-N1 - Mammalian Expression SBFP2 380 446 13 Monomer (A206K) pSBFP2-C1 - Mammalian Expression Azurite...Monomer (A206K) mAzurite-N1 - Mammalian Expression mAzurite-C1 - Mammalian Expression EBFP2 383 448 17 ...

...gRNA_Cloning Vector 41824 Mammalian AfIII none S. pyogenes Bleocin Church MLM3636 43860 Mammalian BsmBI none S. pyogenes...pSPgRNA 47108 Mammalian BbsI none S. pyogenes Gersbach pUC57-sgRNA expression vector 51132 Mammalian BsaI none...sgExpression 48240 Mammalian BbsI yes, activate S. pyogenes Jaenisch pSQT1313 53370 Mammalian BsmBI none S....Gen) 48140 Mammalian BbsI yes, nick S. pyogenes EGFP Zhang PX460 (3rd Gen) 48873 Mammalian BbsI yes, nick... Gen) 48138 Mammalian BbsI yes, cut S. pyogenes EGFP Zhang PX335 (2nd Gen) 42335 Mammalian BbsI yes, nick...Gen) 42333 Mammalian BbsI yes, nick S. pyogenes Puro Zhang PX330 (2nd Gen) 42230 Mammalian BbsI yes, cut...42229 Mammalian BbsI yes, cut S. pyogenes Puro Zhang sgRNA(MS2) cloning backbone 61424 Mammalian BbsI ...

...cTnT Mammalian Campbell 89573 pCAG-iCre iCre CAG Mammalian Wong 89575 pCAG-VCre VCre CAG Mammalian Wong... CAG Mammalian Scholer 125574 pCAG-Cre Cre CAG Mammalian Imai 125577 pCAG-iCre iCre CAG Mammalian Imai...p209 pCMV-cre-K Cre-K CMV Mammalian Green 8395 p210 pCMV-CREM CREM CMV Mammalian Green 8401 p224 pCMV-RFP...floxed RFP within CREM CMV Mammalian Green 8403 p153 pPr-cre Cre probasin Mammalian Green 8404 p236 pPr-CREM...CREM probasin Mammalian Green 8405 p240 pPr-CREM/Ins CREM with insulators probasin Mammalian Green 8406 ...separated by insulator probasin Mammalian Green 11543 pPGK-Cre-bpA Cre PGK Mammalian Rajewsky 11916 pBS185 CMV-Cre...CMV-Cre Cre hCMV Mammalian Sauer 11918 pBS513 EF1alpha-cre Cre EF-1 alpha Mammalian Sauer 11919 pBS448...

...(Anl) Mammalian ATG David Tirrell 64915 pMAH-POLY tyrosyl-tRNA synthetase E. coli pBof Mammalian TAG Huiwang...AbK Mammalian TAG Simon Elsaesser 140011 pAS_4xMx PylT_FLAG-Mx PylRS PylRS M. alvus CpK Mammalian TAG ...-aminotyrosine Mammalian Huiwang Ai 154762 pAS_FLAG-Mma PylRS PylRS M. mazei Mammalian TAG Simon Elsaesser...archaeon Mammalian TAG Simon Elsaesser 154764 pAS_FLAG-Mx1201 PylRS PylRS M. alvus Mammalian TAG Simon...acetyl-lysine Mammalian Tao Liu 164081 pCMV-MfPylRS (human-opti) PylRS M. flavescens acetyl-lysine Mammalian Tao...mazei Mammalian TAG Simon Elsaesser 174896 pAS_4xMmaPylT(UUA)_EF1_MmaPylRS PylRS M. mazei Mammalian TAA ...mazei Mammalian TGA Simon Elsaesser 174901 pAS_MmaPylT_EF1_NES-MmaPylRS WT PylRS M. mazei Mammalian TAG ...

...8729 1859 Mammalian (pcDNA3) GEF Psd 5662 1024 Mammalian (pEGFP-C1) GEF Psd2 84249 771 Mammalian (pEGFP-...Bacterial (pET), Mammalian (pcDNA3), Gateway GTPase Arf3 377 181 Bacterial (pET), Mammalian (pcDNA3), Gateway...Bacterial (pET), Mammalian (pcDNA3), Gateway GTPase Arf5 381 180 Bacterial (pET), Mammalian (pcDNA3), Gateway...Bacterial (pET), Mammalian (pcDNA3), Gateway GTPase Arl1 400 181 Bacterial (pET), Mammalian (pEGFP-N3) Gateway...Bacterial (pET), Mammalian (pcDNA3), Gateway GTPase Arl3 403 182 Bacterial (pET), Mammalian (pcDNA3), Gateway...Bacterial (pET), Mammalian (pcDNA3) GAP Arfgap2 84364 521 Bacterial (pET), Mammalian (pcDNA3) GAP Arfgap3...1704 GAP Arap3 64411 1544 Mammalian (pEGFP-C2) GAP Acap1 9744 740 Mammalian (pFLAG-CMV2) GAP Acap2 23527...

...Backbones Mammalian CMV, SV40, EF1a, CAG, Ubc Transient expression - Gradia Lab plasmids for mammalian expression...or miniTurbo in the mammalian nucleus pSF3-ultraID - Expresses ultraID in mammalian cells with the Tet-...) pcDNA3.1-mCherry - Expresses mCherry tag in mammalian cells pBI-MCS-EGFP - Tet-inducible...FLAG-HA-pcDNA3.1 - C-terminal Flag or N-terminal FLAG-HA for mammalian expression pNIC-CTHF - C-terminal ...HA Epitope tag pcDNA3.1-HA or c-Flag pcDNA3 - Mammalian expression vector with N- or C-terminal HA tag...Myc Epitope tag pKMyc - N-terminal Myc tag for mammalian expression pGEX-4T-1-3xMyc - Bacterial vector ...pEZYmyc-His - C-terminal Myc-His tag for mammalian expression (Gateway) pYIC - Bicistronic fluorescent...

...pAquaN1 Cyan Mammalian Expresses mammalian optimized Aquamarine mCerulean N1 Cyan Mammalian Express a gene...subcellular compartments pCEP4CyPet-MAMM Cyan Mammalian Expresses mammalian optimized CyPet pCyPet-His Cyan Bacterial...Yellow Mammalian Expresses mammalian optimized YPet pYPet-His Yellow Bacterial Expresses YPet with C-terminal...Yellow Mammalian Express a gene of interest fused to the C-terminus of SYFP2 Clover Green Mammalian Expresses...Pre-constructed fluorescent biosensors targeting small molecules or specific genes are also available. ...N-terminus of monomeric Cerulean mCerulean C1 Cyan Mammalian Express a gene of interest fused to the C-terminus...C-terminus of monomeric Cerulean mTurquoise2 Cyan Mammalian Constructs to target mTurquoise2 to various subcellular...

...promoter for normalization. FMDV StopGo sequences flank the MCS for improved normalization. John Atkins... CMV Mammalian expression of Nano-lantern Takeharu Nagai 87121 pcDNA-RLuc8 Renilla CMV Mammalian expression...of organisms, ranging from bacteria to aquatic animals to insects. Luciferase catalyzes the oxidation ...that has been codon optimized for expression in mammalian systems. Other luciferases in our collection include...has also been codon optimized ( CBR2opt ) for mammalian expression systems. Each type of luciferase has...due to their large size. NanoLuc® Luciferase is small, bright, and a secreted version of the protein is...protein interactions , protein dimerization , endosomal escape of cell-penetrating peptides , and more...

...Browse Plasmids: Anti-CRISPR Small Precision Edits CRISPR can be used to make small, very precise edits of ...double-stranded DNA plasmid. Small ssDNA oligos often work well for small edits like a single-point mutation...Guide Jump To: CRISPR Guide Overview CRISPR Basics Small Edits Large Edits Genome-Wide Screening Cas9 Fusion...missing sequence, and is more commonly used for small precision edits. NHEJ directly ligates the break...efficient in repairing DSB, it frequently causes small nucleotide insertions or deletions (indels) at the...modification ). In most cases, NHEJ gives rise to small indels in the target DNA that result in amino acid... handful of bases, or even a single base. These small precision edits can be done via HDR, base editing...

...methods: in animals, in cultured immune cells, or through the use of recombinant plasmids. Animals - Antibodies...Antibodies produced in animals make use of the animal’s immune system. The antigen of interest is introduced...introduced to the animal, which then mounts an immune response to it. Once the immune response has developed...antibody. Monocultures of B cells isolated from animals immunized as described above are used to create... protein chains: two large heavy chains and two small light chains. These four domains come together to...only produce one variable region. Isotypes The mammalian immune system produces five classes of antibodies...antibodies are typically collected directly from animals and contain antibodies targeting various epitopes...