...Protocols ddPCR Titration of AAV Vectors ddPCR Titration of AAV Vectors You may also like... AAV Titration...Titration by qPCR AAV Viral Service Viral Vector Guides Virus Blog Posts Molecular Biology Reference Introduction...titer purified recombinant Adeno-associated viral vectors (AAV). This protocol specifically uses primers ...probes targeting the ITR elements in the viral vectors but can be modified for other targets. The dilution...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette 1–10 µL multichannel pipette...calculate the titer for each sample: $$T = {R*C*D*1000\over V}$$ Where: T = Titer, GC/mL R = Reaction volume...

...Lentivirus Production You may also like... Viral Vector Guides Mol Bio Protocols Introduction This protocol...be used to produce lentivirus from a lentiviral vector transfected into 293T cells using a polyethylenimine...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Ice bucket CO 2 incubator...solution through a 0.22 μm membrane. Aliquot 500–1000 μL into sterile tubes. Store the tubes at -20 °C...PEI such that the ratio of μg DNA:μg PEI is 1:3 (1000 μL total per 10 cm dish). Using transfer plasmid...

... gene of interest from an integrated lentiviral vector....Lines with Lentivirus You may also like... Viral Vector Guides Virus Blog Posts Mol Bio Protocols Viral... gene of interest from an integrated lentiviral vector. Unlike the short-term protein expression observed...approaches, generating cell lines using lentiviral vectors enables long-term protein expression studies. Moreover...repeated transient transfection. Some lentiviral vectors deliver mammalian antibiotic resistance (e.g., ...lentiviral transgene. Note that not all lentiviral vectors deliver antibiotic resistance. This protocol was...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Ice bucket CO 2 incubator...

...Assay for Lentivirus You may also like... Viral Vector Guides Virus Blog Posts Mol Bio Protocols Viral...marker. Determining the titer of your lentiviral vector allows you to control the multiplicity of infection...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Ice bucket CO 2 incubator...protocol was developed using low titer lentiviral vectors (10 5 TU/mL). If you anticipate that your viral...

...by Limiting Dilution You may also like... Viral Vector Guides Virus Blog Posts Addgene Protocols Viral...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette CO 2 incubator Pipet controller...96-well plate, seed one of the corner wells with ~1000 cells. This will help you focus the microscope later... incubating the cells. Pro-Tip Use the well with 1000 cells to focus the microscope, then you can easily... plasmid #52962 ) and is described in Improved vectors and genome-wide libraries for CRISPR screening....

...Assay for Lentivirus You may also like... Viral Vector Guides Virus Blog Posts Mol Bio Protocols Viral...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Ice bucket CO 2 incubator...the indicated serial dilutions of the lentiviral vector pRosetta . The following day, cells were treated...the indicated serial dilutions of the lentiviral vector pHAGE EF1α dCas9-KRAB . The following day, cells...

...General Transfection You may also like... Viral Vector Guides Virus Blog Posts Mol Bio Protocols Viral...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Ice bucket CO 2 incubator...solution through a 0.22 µm membrane. Aliquot 500–1000 µL into sterile tubes. Store the tubes at -80 °C...

...ddPCR) for lentivirus vector titration.... Protocols ddPCR Titration of Lentivirus Vectors ddPCR Titration of...of Lentivirus Vectors You may also like... Lentivirus Colony Formation Titration Assay Lentivirus Fluorescence...droplet digital PCR (ddPCR) to titer lentivirus vectors. This protocol specifically uses primers and probes...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette 2–50 µL multichannel pipette...

...Cells You may also like... AAV Viral Service Viral Vector Guides Virus Blog Posts Molecular Biology Reference...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette Hemocytometer or cell ...system, Nalgene, 565-0010 (or for larger volumes 1000 mL 0.45 µm Rapid-Flow PES Filtration Unit, Nalgene...solution through a 0.22 μm membrane. Aliquot 500–1000 μL into sterile tubes. Store the tubes at -80 °C... deionized water and adjust to a final volume of 1000 mL. Stir at RT until fully dissolved. Pro-Tip This...

...approximately 10-fold lower for ligation of inserts to vectors than for an intact control plasmid. Incubate the...when using highly competent cells. If you used 100-1000 ng of total DNA in a ligation you will often get...

...SYBR Green Technology You may also like... Viral Vector Guides Virus Blog Posts Mol Bio Protocols Introduction...the titer of samples obtained from academic viral vector cores. Our titers were similar to those reported...channel pipette 20–200 µL single channel pipette 200–1000 µL single channel pipette 1–10 µL multichannel pipette...95 uL 20X 200X Dilution 3 20 uL Dil. 2 80 uL 5X 1000X Dilution 4 20 uL Dil. 3 80 uL 5X 5000X Dilution ...

... Ultracentrifugation You may also like... Viral Vector Guides Virus Blog Posts Mol Bio Protocols Introduction...and characterization of adeno-associated viral vectors," (Link opens in a new window) copyright (2006)...bring up the volume with formulation buffer. Use a P1000 to the bottom of the filter and pipette up/down ...