...proceeds. One popular lox-stop-lox plasmid is from Tyler Jack's lab: Lox-Stop-Lox TOPO.  Cre-dependent ...tissue-specific cell types: Cre-lox recombination. What is Cre-lox? The Cre-lox system is a technology that...translocation event is generated at the loxP sites. How can I use Cre-lox? The Cre/lox system is a well-established...termination features with loxP sites on either side (often called a “lox-stop-lox” or “LSL” cassette) upstream...Among these are loxN (GtATACcT), lox2272 (GgATACtT), and lox511 (GtATACAT). These variant lox sites undergo...Cre and loxP, respectively. The FLP-FRT technology can be an effective alternative to Cre-lox, and has...embryonic lethality. To facilitate the use of Cre-lox technology, transgenic mice have been constructed...

...have two pairs of lox sites which are part of a dual lox system called FLEX or DIO. Lox sites are recognized...transfer plasmid, but are often seen in them.  Lox sites Lox sites, in combination with the Cre recombinase...spatially and temporally control transgene expression. Lox sites are directional 34 bp sequences that can flank...first flips the sequence between the first pair of lox sites, and then excise the sequence between the second...transgene. By restricting the availability of Cre, lox sites lets researchers control the spatial and temporal...activation of a transgene. Figure 2: FLEX Cre-Lox system.  WPRE The Woodchuck Hepatitis Virus (WHP...

...continued utility of Cre-lox. Check out our previous blog post, Plasmids 101: Cre-lox, if you need a quick...quick primer on how Cre-lox recombination works. Advances in Cre-lox recombination have included inducible...manipulation systems remain valuable to biology. Cre-lox recombination, discovered in the 1980s, is one of...expression, especially in in vivo models, and new Cre-lox based technologies are still being developed today...tamoxifen-responsive CreERT, the coupling of Cre-lox with FLP-FRT recombination to generate additional...Cellular Biology, harnessed these attributes of Cre-lox to create the Brainbow mouse neural labeling system... cre-lox...

...heterozygous for each mutated allele.  Cre-lox expressing strains Cre-lox strains are a common example of mice...Cre-lox Read our genome engineering blog posts Resources on Addgene.org Find more about Cre-lox Read...expressing both Cre and LoxP occurs. But how are these mice made to begin with? Cre-lox mice are made from ...cross mice to create double knockout lines and Cre-lox lines, and how to properly control for genetically...recombination at the floxed site, your control should be littermates that express Cre but not loxP. With this two-part...that expresses Cre with a strain that expresses a floxed gene of interest. Cre may be expressed globally... is a dominant phenotype, while you’ll want the floxed gene to be homozygous in order for Cre-mediated...

...Cre-Lox Check out Retrograde AAVs for Neurscience Research Resources on Addgene.org Find Cre-Lox Plasmids... in a similar fashion as between loxP sites (For details see Cre-lox blog post). Optimizing FLP FRT through... use the Cre-lox or one of the FLP-FRT systems in your experiments. Mutated FRT and loxP sites in Flex... contributed by guest blogger Katrin Michel. Cre-lox is an incredibly popular and powerful site specific...developed. Read on to learn more about FLP-FRT, Cre-lox, and how combinations of FLP and Cre enable additional... cre-lox...recombination between those sites. FRT sites and loxP sites differ at the nucleotide level but share an...

...recombinase, which induces recombination at LoxP sites. Placing a lox-STOP-lox cassette upstream of a gene of interest...T-DDOGs can be used in combination with green-red Cre-lox systems. T-DDOGs can also be designed with other ...121. PubMed PMID: 22157958. Addgene Resources Cre-lox Resource Guide Fluorescent Protein Resource Guide... cre-lox...

... recombinase-dependent vector designs used a lox-STOP-lox (LSL) or frt-STOP-frt (FSF) sequence prior to...unused (loxp) sites will be flanked by two lox sites now in the same orientation. Next, the loxp site is... to the system it binds one pair of lox sites, let's say the loxp pair, and flips the transgene into the...Cre-recombinase binds to either pair of lox sites, let's say the lox2272, that are in the opposite orientation...Recombinase-dependent vectors utilize pairs of specific sequences, lox (recognized by Cre) or frt (recognized by Flp), to... hypothesis, they randomly scrambled one pair of lox sites and either used the same sequence to flank ... in one loxp site and one lox2272 site on either end of the transgene. Since loxp and lox2272 can not ...

...Cre-lox Read our genome engineering blog posts Resources on Addgene.org Find more about Cre-lox Read..., conditional knockout/knockin mice, use the Cre-lox recombination system to express the transgene in ... expression of the target gene. In addition, Cre-lox systems can be used to invert a segment of DNA or...recombination of DNA sequences located between two loxP sites. The loxP sites are present at both ends of a gene... vector, creating the floxed gene segment. Crossing of mice containing a floxed gene to a Cre-expressing...recombination. To generate a conditional knockout mouse, loxP sites are inserted directly into the genome to flank...gene in the targeting vector. The stop cassette is floxed, so Cre-mediated recombination deletes the stop...

...Learn more about Mammalian shRNA Tools for RNAi! Cre-Lox technology CAG-GFP-IRES-CRE (Plasmid #48201) This...Gage lab encodes GFP and Cre-recombinase. The Cre-lox system is a technology that can be used to induce... to use the pLKO.1 vector can be found here. Cre-Lox technology Puro.Cre empty vector (Plasmid #17408)...

... after selection using the Cre/Lox recombination system. After “floxing” the resistance gene through the...conditional knockout, researchers often use the Cre/Lox system described earlier. In this case, you design....org Find Mammalian Expression Vectors Find Cre-Lox Plasmids Learn More about CRISPR  ...NeoR only and leave exon 2 floxed, as seen in line 4 of Figure 4. Since the loxP sites are located in intronic...template. These templates can include loxP sites to create conditional floxed alleles. Although CRISPR is very... the insertion of flanking LoxP sequences, you can remove the gene through the addition of Cre recombinase...this construct, the neomycin resistance gene is floxed so that it can be removed post-selection through...

...Cre-lox recombination is an incredibly useful molecular biology tool, but like any biological system,...  cre-lox...Cre may induce recombination at pseudo- or cryptic loxP sites (estimated to occur at a frequency of 1.2 ... systems, Cre itself, without the presence of a floxed construct, may produce a phenotype. This problem...458-65. doi:10.1007/s004270100167. PubMed. Cryptic loxP sites in mammalian genomes: genome-wide distribution...

...application of iTOP Having tested iTOP using Cre-lox recombination, D’Astolfo et al. were eager to see... cre-lox...series of primary cell types using Cre and various loxP reporters. In both mouse and human embryonic stem...

...   Recombinases Recombinase systems such as Cre-lox allow for more accurate cell and tissue targeting...functional Cre enzyme that induces recombination of lox sites. This system takes advantage of the extensive...Deisseroth 112677 AAV2 pOTTC1032 - pAAV EF1a Nuc-flox(mCherry)-EGFP Brandon Harvey 50457 AAV2 pAAV-hSyn-DIO-EGFP...

...Plasmids 101: The Promoter Region Plasmids 101: Cre-lox Light-Switchable Two-Component Systems for E. coli...Tetracycline (Tet) On Expression Systems at Addgene Cre-lox Guide and Plasmids     ...

...and reduces toxicity as compared to previous Cre/lox or drug-induced Cre systems (ex: tamoxifen). As the... the name suggests, CreLite combines the Cre/lox system with optogenetics. Cre activity is controlled ...

...bacteriophage integrase phiC31. The popular UAS/Gal4 and Cre/Lox gene regulatory systems have also been successfully...zebrafish transgenesis with Tol2 and application for Cre/lox recombination experiments." Methods in cell biology...

...using Cre-Lox mouse models in which the endogenous MECP2 gene was disrupted and silenced by a Lox-Stop cassette...

...presence induces first opsin inversion, and then lox sites excision, therefore locking the opsin into ... Visit our molecular biology reference Find Cre-lox plasmids at Addgene Browse plasmids at Addgene ...is flanked (“floxed”) by two sets of incompatible recognition sites (e.g., loxP and lox511) in opposite...flip-excision) switch (aka DIO (Double-floxed Inverse Orientation) or DO (Double-floxed Orientation)) is a powerful...recombination at defined recognition sites (i.e., loxP and FRT, respectively) (Abremski and Hoess, 1984...however, Cre induces the recombination of the double-floxed opsin construct, thereby enabling its expression... Van Duyne, Gregory D. "A structural view of cre-loxp site-specific recombination." Annual review of biophysics...

...system for controlling gene expression such as Cre-lox, may be sufficient to overcome the transport issues...to produce neuron specific transfection using Cre-lox, this new method could allow for labeling of neurons...

...in CRISPR genome editing.   2. Plasmids 101: Cre-lox Cre recombinase can be used to generate inversions...

... Cre/lox recombination system. A researcher can flank fluorescent proteins in a plasmid with loxP sites...respectively. The permanent labelling of cells using cre-lox is advantageous when compared to photoactivable or...

...Resources on the Addgene Blog Advanced Uses of Cre-lox and Flp-FRT - A Neuroscientist's View Interview: ...

...expression systems exist (eg. LexA/LexAop and Cre/Lox systems) in case you want to express one gene in ...

...good, the Cre-Lox system offers inducible yet permanent gene excision. Essentially, LoxP sites are knocked...time-tested system but does require knock-in of the LoxP sites, which is more difficult than a simple knock...out plasmids? Check out our Cre vector and Murine LoxP vector.   Turn it off for a little while Transient...

...Fluorescent Proteins Control Protein Expression with Cre-lox Control Protein Activity with GFP Resources at Addgene.org...

...lentiviral, retroviral, AAV, RNAi, luciferase, cre/lox, yeast expression, worm expression, insect expression...

...Engineering Guide CRISPR Mammalian Expression plasmids Cre-Lox plasmids References Prathipati, P., Rodriguez-Aguayo...

...Delivery to the CNS and PNS Advanced Uses of Cre-lox and Flp-FRT - A Neuroscientist's View Control Gene...

...Specific Recombination in Drosophila Learn about Cre-Lox Resources on Addgene.org Find Drosophila Plamids...

...orange) (Livet et al., 2007). Brainbow uses a Cre/lox system to label neurons. Genes encoding the fluorescent...Brainbow 2.1 construct in the Rosa26 locus, under a loxP “roadblock” so that XFP are only expressed following...

... safe for clinical applications. Excisable Cre-lox lentiviral vectors: This approach is the same as ...transgenes that are flanked by loxP sites. While there is only a small loxP scar behind at the site of excision...

... involving recombination elements such as FRT or Lox sites, and provides a modular system for easier exchange...recombination events at the genetic level. Upon Cre/loxP recombination, each transgene expresses one of three...recombinase/FRT system in place of Brainbow’s Cre/loxP, allowing for simultaneous use of the two systems...

...Both loxP and lox511 are recognized by Cre but lox511 sites can only recombine with other lox511 sites... the identical loxP or lox511 sites (Figure 1, bottom). Since only one loxP and lox511 site will remain...with one loxP site and one lox511 site in the same orientation, and placing the additional loxP and lox511...binds loxP sites, while FLP binds FRT sites. When a DNA sequence is flanked by target sites (floxed) in ...one wild-type pair (loxP, black arrowheads) and one mutated pair (for example- lox511, white arrowheads)...sites, not with loxP sites. Alternatively, you could use a pair of loxP sites and a pair of FRT sites and... proceed either by first utilizing the loxP sites or the lox551 sites. Regardless, the first recombination...

...allele The design for a floxed conditional allele requires you to target 2 loxP sites to intronic regions...original design for creating a conditional allele, or floxed mouse, from Yang et al. requires two gRNAs targeting... and two repair templates in order to insert two loxP sites. Since two double-strand breaks are made, ...variety of other undesired outcomes, including one loxP site insertion and one NHEJ-repaired indel, often... - 0.8 kb apart. In the example below, we’ll add loxP sites for conditional deletion of exon 6 of Syt1...homology on each side. For optimal repair, place the loxP sites precisely at the cut site for each guide. ...efficiency of Easi-CRISPR for simple knock-ins and floxed models varies from 8.5-100% of pups born, with ...

...technology (Cre/loxP) to remove the donor cassette. While this method can be effective, a loxP site is left...Grégoire and M. Kmita, “Recombination between inverted loxP sites is cytotoxic for proliferating cells and provides...

...pAAV-mDlx-NLS-mRuby2 112677 AAV1, AAV2 pOTTC1032 - pAAV EF1a Nuc-flox(mCherry)-EGFP 27056 AAVrg pAAV-Ef1a-DIO EYFP ... mCherry 112677 AAV2 pOTTC1032 - pAAV EF1a Nuc-flox(mCherry)-EGFP 50457 AAV2 pAAV-hSyn-DIO-EGFP ...

...easy to remove after injection, they added flanking loxP sites and a heat-shock inducible Cre to generate...schematic below.) After self-excision, the resulting loxP scar is located in this synthetic intron, and only...

...GABAergic interneurons. pOTTC1032 - pAAV EF1a Nuc-flox(mCherry)-EGFP (112677-AAV5 and 112677-AAVrg): The...)-mCherry (26976-AAV5) pOTTC1032 - pAAV EF1a Nuc-flox(mCherry)-EGFP (112677-AAV5) Calcium Sensors pGP-AAV-syn-jGCaMP7b-WPRE...

... of gene-specific 1 kb homology arms flanking a FLOXed neoR gene. pAAV-SEPT-Acceptor, the preferred vector...Then, if necessary for subsequent experiments, the floxed neoR gene is removed via infection with adeno-cre...

...Lab used this backbone to target a CAG-driven loxP-stop-loxP cassette into the Rosa26 locus, allowing for...Commonly used sequences, including fluorescent tags, a floxed Cbr-unc-119 selectable marker, and “connectors”...

...Combinations of classical genetic tools, for example Cre/loxP or Flp/FRT, with AAV vectors are common approaches...

...stochastic multicolor labeling, Brainbow used the Cre-loxP system to express one of the three fluorescent protein...

...pAAV-hSyn-DIO-hM4D(Gi)-mCherry contains a double floxed human muscarinic receptor 4 (Gαi-coupled) DREADD...

...popular method uses Cre recombinase on transgenic flox animals to introduce modifications at the DNA level...

...these needs is the lentiviral-based expression of floxed dCas9 so that it may later be excised by Cre recombinase...

...N. K., Lee, C. K., Abu Bakar, S., AbuBakar, S., Hollox, E. J., & Boon Peng, H. (2015). A comparison of...

...nonfluorescent (Halo, SNAP) tags, a selectable marker (floxed Cbr-unc-119) for easy screening of the insertion...

...the cap gene. In this special CREATE cap plasmid, loxP sites flank the downstream polyA sequence. When ...

...-made AAV vectors which can be used in numerous floxed animal species for behavioral tests.    Forced ...

...(1979) Transfer of proteins from gels to diazobenzyloxymethyl-paper and detection with antisera: a method...