We narrowed to 965 results for: REP

...conditioned medium prepared above to make a new cell solution at a concentration of 5 cells/mL. Prepare approximately...culture dish, Corning 430167 (optional) Reagent Preparation DMEM Complete: 10% v/v FBS and 4 mM L-alanyl-...well plate. Allow the cells to grow overnight. Replace the medium on the cells and collect the used (conditioned...a hemocytometer or other cell counter. Pro-Tip Prepare a relatively dilute cell solution (<10 6 cells/.... If you choose not to use conditioned medium, prepare this cell solution in fresh complete medium. This...

...concentration ( see Preparing Antibiotics ). Note: If you intend to do a miniprep, you will usually want...inoculate bacteria in liquid culture. Written Protocol Prepare liquid LB. For example, to make 400 mL of LB, weigh...want to start 2 mL in a Falcon tube. For larger preps, you might want to use as much as a liter of LB ...without growth (top) and with growth (bottom). Preparing Antibiotics Create a stock solution of your antibiotic...

...2 (am): 18 h post transfection - Remove media, replace with fresh media Day 3 or more (am): Observe fluorescence... (PES) filter Syringes for filtering Reagent Preparation DMEM Complete: 10% v/v FBS and 4 mM L-alanyl-...empirically determined for each new batch of 1 mg/mL PEI prepared. There may be variation between batches of PEI...empirically determined. Once a batch of PEI is prepared, transfect cells with a fluorescent plasmid using...following morning, carefully aspirate the media. Replace the media with 15 mL of DMEM complete. Incubate...

...the full details of our protocols supports reproducibility and accelerates science. Here, we list the ...solely for transparency intended to support reproducibility in science. Last Update: January 20, 2022 Workflow...primary antibodies raised in a mouse. Reagent Preparation Permeabilization buffer: Dilute 20 µL of Triton...100 in 50 mL PBS. 300 nM DAPI working solution: Prepare a 300 µM DAPI stock solution by diluting 2.1 µL...DAPI solution to 100 µL PBS. Protect from light. Prepare a 300 nM DAPI working solution by diluting 5 µL...

...15-25% glycerol. You can prepare the glycerol stock the same time you prepare your plasmid DNA. In the...able to start an overnight culture for plasmid DNA prep the following day. Tips and FAQ The optimal concentration...glycerol stock before you begin your plasmid mini-prep. Try not to freeze/thaw your glycerol stock too ...

...result represents a single experiment. For publishable data, the experiment would need to be repeated to ...their resistance genes, as one does not need to prepare separate batches of antibiotic-containing agar....or other antibiotic resistance) Procedure Day 1 Prepare carbenicillin to a concentration of 1 mg/mL – 4...

...bottom. For the P1000, this represents the ones. For the P200, this represents the first decimal place. ...For the P20, this represents the second decimal place, and for the P2 this represents the third decimal...

... but nicked vector product is then repaired during the replication cycle. Empty vectors for LIC typically... plasmid together through the transformation/replication process. LIC employs long overhangs to form a...

...Purification Purify adeno-associated virus from a preparation produced in mammalian cells AAV Purification ... Inoculating a Liquid Bacterial Culture How to prepare and grow bacteria in a liquid medium Inoculating...physical titer of your adeno-associated virus preparation. AAV Titration by qPCR Using SYBR Green Technology...

...bacteria are used as the means for both storing and replicating plasmids. Because of this, nearly all plasmids...expression) carry both a bacterial origin of replication and an antibiotic resistance gene for use as ...sell competent cells, which come frozen and are prepared for optimal transformation efficiencies upon thawing...

...the new target DNA strand accurately and rapidly. Repeat steps 2-4 25-30 times. Final Extension for 5 minutes...°C below Tm). Extend DNA for 2 minutes at 72°C. Repeat steps 8-10 for 25-30 cycles. Final Extension for...and concentration of PCR product. Master Mix Preparation Multiply the volume of each reagent by the number...

...table, reprogramming efficiencies ranking as low, medium or high is based on the reprogramming efficiencies... be used to package reprogramming factors. An important milestone of reprogramming is the silencing of...incomplete silencing of reprogramming transgenes, which leads to incomplete reprogramming. Additionally, lingering...rates of reprogramming when compared to MMLV-derived retroviruses, it is not as well repressed following...by terminal repeat sequences, which the PB transposase recognizes. Following reprogramming, transgenes...risk of lingering expression of reprogramming factors. Non-replicating Plasmids: Generating iPSCs with...RNA to reprogram cells is that it’s simple and efficient. It has a high efficiency of reprogramming when...

...Origin of Replication? See our Plasmids 101: Origin of Replication blog).  The replication problem Plasmids...directly inhibit replication by binding to the origin of replication and blocking replication machinery (del...inhibit plasmid replication.   Figure 1: The replication problem. Iterons are repeated sequences within...increases, RepA transcription is repressed and replication is downregulated. In the second model, termed...cell have the same replicon and/or partitioning system. Let’s start with the replicon- the part of the ...plasmid that contains the origin of replication and the replication control machinery (Need a refresher... with the same replicon are incompatible because they compete for the same replication control machinery...

...but browsing the 49 new preps available in the Addgene viral vector repository with a nice cup of hot ...viral prep with multiple serotypes pAAV-hSyn-GRAB_OTmut AAV1 Biosensor Yulong Li New viral prep pAAV-CAG-NIR-GECO2G...Fyhn New viral prep AAV-hSyn-Ribo-jGCaMP8m AAV1 Biosensor Marianne Fyhn New viral prep pAAV-Ef1a-fDIO... viral prep pAAV_BiSSTe10_dTomato_nlsdTomato AAV-PHPeB Control Gordon Fishell New viral prep pAAV_... viral prep pAAV_BiCHATe27_dTomato_nlsdTomato AAV-PHPeB Control Gordon Fishell New viral prep pAAV_BiCHATe27...New viral prep pAAV_BiVIPe4_dTomato_nlsdTomato AAV-PHPeB Control Gordon Fishell New viral prep pAAV_... viral prep pAAV_BiLAMP5e3_ChR2_mCherry AAV-PHPeB Optogenetics Gordon Fishell New viral prep pAAV_...

...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...

...been designed by the Addgene nonprofit plasmid repository and is being shared outside of Addgene for informational...informational purposes only. If you choose to reuse or repurpose this SOP in another location, please note that...

...fused to a streptavidin core that are able to retain reporter proteins fused to the streptavidin-binding ... the 3’ UTR of the reporter mRNA via an MS2 coat protein (NLS-MS2-RFP) the reporter mRNA, which contains...their reporter mRNAs. Using the TRICK plasmids, Halstead et al demonstrated that 91% of reporter mRNAs...customized fluorescent reporters can be challenging enough, but incorporating multiple reporters into a single ...pyralis) reporter plasmids for use in high-throughput screening of pre-mRNA splicing. This reporter system...activation or repression. The TCS light sensors described in ACS Synthetic Biology (2014) represent the newest... from the reporter construct. The sgRNA targeting Gaussia luciferase can be easily replaced with an sgRNA...

...and reprogramming and direct reprogramming Explore different methods for delivery of reprogramming factors...prior to reprogramming (CellTagMEF), at the end of the three-day transcription factor reprogramming treatment...Direct reprogramming describes the process where differentiated cells are turned into a cell type of ...valuable tool for regenerative medicine, direct reprogramming is an inefficient process, with the majority...patterns unique to the few cells that successfully reprogram. Unfortunately scRNA-seq only captures the gene...investigate how early expression patterns influence reprogramming outcomes. CellTag from the Morris Lab at Washington...linking ancestral gene expression patterns with reprogramming outcomes (Biddy et al., 2018). Tracking lineage...

... new preps available in the Addgene viral vector repository! This includes the first Addgene prep packaged...viral prep pAAV_hSyn-DIO-PdCO-mScarlet-WPRE AAV1, AAV5 Optogenetics Ofer Yizhar New viral prep with...Lin Tian New viral prep pAAV-hSyn-GRAB-gDA3m AAV1 Biosensors Yulong Li New viral prep pGP-AAV-GFAP-iGABASnFR2...part of the BRAIN Armamentarium Collection. These preps drive expression of either Cre recombinase or a ...New ready-to-use viral vectors in the Addgene repository  Name Serotype Category Depositor  Notes ...Recombinases Stanley Thomas Carmichael New viral prep pAAV-EF1a-fDIO-Cre AAV5 Recombinases Esteban ... AAV PHP.eB Recombinases Jonathan Ting New viral prep AiP13779 - pAAV-AiE0779m_3xC2-minBG-iCre(R297T...

...required for replication and transcription (Figure 1). All viral genes are removed and replaced by a single...signals necessary to direct minigenome replication and reporter gene transcription by the EBOV polymerase...Technical Tips). Minigenome replication and transcription In order to replicate the minigenome, the viral...transcription and replication strategies of Marburg virus and Ebola virus by using artificial replication systems...allowing researchers to study aspects of the EBOV replication cycle under BSL-2 conditions (4). What is a ...that are crucial cis-acting signals for genome replication and viral gene transcription (6). The EBOV genome...single (non-viral) reporter gene (e.g. firefly luciferase or eGFP) flanked by the EBOV-specific gene start...