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..., the nucleotides on the opposing strands of DNA form hydrogen bonds, where A always pairs with T, and...broadly in their functionality. In their simplest form, plasmids require a bacterial origin of replication...plasmid. The selectable marker is typically in the form of another antibiotic resistance gene under the ...fluorescence detection, and the sequence outputted in the form of a chromatogram. Sanger can sequence approximately...common is the Illumina NGS platform for short-read sequencing. This is the platform Addgene primarily uses...entire sequence using bioinformatic methods. It is important to note that NGS platforms in general do not ...The concept of genes as carriers of phenotypic information was introduced in the early 19th century by Gregor...

...your purified virus. Nonetheless, we recommend performing a buffer exchange before using the purified AAV...0.001% Poloxamer 188 in PBS + 200 mM NaCl (C) (formulation buffer) Add 5 mL of Buffer B and 2 mL of 5 M ...supernatant on top of the gradient. Use 1X PBS (or formulation buffer) to top off the tube. Seal the QuickSeal...not easily removed. After each spin, add more formulation buffer and sample and make sure to pipet back...is less than 500 µL, bring up the volume with formulation buffer. Use a P1000 to the bottom of the filter...

...four biosafety levels. This protocol provides information for both biosafety level 1 (BSL-1) and biosafety...to your hand, but not so tight that you cannot perform your duties. Be sure to try out different sizes... and is being shared outside of Addgene for informational purposes only. If you choose to reuse or repurpose... can be held responsible for your use of the information contained in or linked to from these web pages...

... tips Bacterial Transformations We walk you through a standard plasmid transformation protocol and offer...tricks to reduce transformation time and increase efficiency Bacterial Transformation Protocol Inoculating...

...four biosafety levels. This protocol provides information for both biosafety level 1 (BSL-1) and biosafety...clearly marked as “BSL-2.” The names and contact information of the laboratory manager should be clearly visible... and is being shared outside of Addgene for informational purposes only. If you choose to reuse or repurpose... can be held responsible for your use of the information contained in or linked to from these web pages...

... are multiple objective lenses. Each objective performs essentially the same task but has different magnifying...an objective, you will see several pieces of information, but the most important (for now) is the magnification..., any stains used in your sample, and sample information so that you or others can interpret your images...scratches, and dust can easily impair microscope performance. Be sure to clean up your work area and microscope...

...specimens. These technologies have the potential to transform neuroscience research by bringing the precision...

...Digest of Plasmid DNA DNA Ligation Bacterial Transformation Summary Oligo overlap cloning can be used anytime...that can be added when ordering them or can be performed enzymatically later. Experimental Procedure Digest...the vector with 0.75-6 ng of annealed oligos). Transform 2-3μL into your favorite competent bacteria and...

...Promoter Plasmids using the table below. Additional information is available for each construct and can be accessed...

...light chains. These four domains come together to form a “Y” shape, as shown in Figure 1. The two arms ...the five human isotypes. Some isotypes naturally form multi-antibody units, while others exist as monomers...may want to identify or collect the protein in any form it may be present in. However, polyclonals can vary...antibody, for each antibody used. Blank controls in the form of sample buffer must also be run. Unique samples...fluorophores. Horseradish peroxidase (HRP) HRP performs a chemiluminescent reaction in the presence of...activated and its output measured. This method can be performed using the indirect detection method via a conjugated... able to recognize the protein in its native conformation. Attachment of the antibody to the beads is ...

... the Video! DNA Purification Miniprep, phenol-chloroform extract, and precipitate DNA DNA Quantification...Ligation Assemble DNA using DNA ligase Bacterial Transformation Introduce DNA into a bacterial strain Watch... virus using fluorescence measurements Colony Formation Titering Assay for Lentivirus Quantify virus by...

...specific products or equipment. Inclusion of this information is solely for transparency intended to support... counter 37 °C, 5% CO 2 incubator with shaking platform set to 120 rpm 37 °C bead bath Vortex Stir bar...flask.Incubate in a 37 °C, 5% CO 2 incubator on a shaking platform set to 120 rpm. Pro-Tip Do not use cells that ...

...specific products or equipment. Inclusion of this information is solely for transparency intended to support...spectrophotometer 37 °C, 5% CO 2 incubator with shaking platform set to 120 rpm 37 °C bead bath Clamp stand and...divide the sample among multiple columns. For information about the total binding capacity for your columns...

...potential consequences of having a non-functional form of your protein expressed in your system. HDR : ... to produce the Cas enzyme, and the enzyme then forms a complex with the gRNA for CRISPR activity. Because...purified Cas protein and in vitro transcribed gRNA to form a Cas-gRNA complex. You then delivery the RNP to... and large insertions or deletions. For more information on the wide range of CRISPR technologies, visit...Selecting the genetic manipulation you plan to perform will help you narrow down which components you’...edit, ideally less than 10 bp away. For more information, read our three tips to improve HDR efficiency...your Cas protein and gRNA, which you generally perform by chemical transfection or electroporation. Expression...

...hort P alindromic R epeat (CRISPR) systems, which form an adaptive immune system in bacteria, have been...genome engineering. Find essential background information on CRISPR and the basics for planning your first...

... Gel Electrophoresis DNA Ligation Background Information Gel purification allows you to isolate and purify...step with the now gel purified DNA. Find more information about DNA quantification here . Tips and FAQ ...

...Protocol, tips, and FAQ for how to perform a restriction digest of plasmid DNA...more enzyme is advisable. Reactions are often performed with 0.2-0.5 µL of enzyme because it is difficult...

...containers Reagents Liquid for pipetting Background Information Anatomy of a Pipette This front view photo of...the Volume on the Pipette Although each pipette performs the same function, the numbers are read differently...

...Standard protocol for performing agarose gel electrophoresis, including tips to improve resolution and...Watch the protocol video below to learn how to perform gel electrophoresis. Equipment Casting tray Well...

...Follow this protocol to perform digital droplet PCR (ddPCR) for lentivirus vector titration....Vectors You may also like... Lentivirus Colony Formation Titration Assay Lentivirus Fluorescence Titration...