![]() This improved assembly protocol provides opportunities to analyze any five desired pathways at once much quicker. Here we present an improved three-step synthetic assembly protocol to quickly and efficiently generate multiplex hextuple luciferase reporter plasmids for other signaling pathways of interest. The same synthetic assembly cloning pipeline allows the stitching of numerous other cellular pathway luciferase reporters. Our proof-of-concept multiplex hextuple luciferase assay was designed to simultaneously monitor the p53, TGF-β, NF-κβ, c-Myc, and MAPK/JNK signaling pathways. Because all six reporters are on a single piece of DNA, a single vector ensures stoichiometric ratios of each transcriptional unit in each transfected cell, resulting in lower experimental variation. We used synthetic assembly cloning ( Vazquez-Vilar, et al., 2020) to assemble all six luciferase reporter units into a single vector, over eight stitching rounds. We recently developed a multiplex luciferase assay that allows monitoring the activity of five experimental pathways against one control simultaneously. They are genetically encoded, versatile, and cost-effective, whose output signals can be sensitively detected. Luciferases are good candidates for generation of such signals. The signals from such measurements should be independently detectable and measure large dynamic ranges. While most cell-based assays measure single quantities, multiplexed assays seek to address these limitations by obtaining multiple simultaneous measurements. Such assays are designed to examine the effects of small compounds on targets, pathways, or phenotypes participating in normal and disease processes. If (-not (Get-Command choco.High-throughput cell-based screening assays are valuable tools in the discovery of chemical probes and therapeutic agents. zip to the filename to handle archive cmdlet limitations # Ensure Chocolatey is installed from your internal repository # $Chocolate圜entralManagementServiceSalt = "servicesalt" # $Chocolate圜entralManagementClientSalt = "clientsalt" # $Chocolate圜entralManagementUrl = " # ii. # If using CCM to manage Chocolatey, add the following: ![]() $ChocolateyDownloadUrl = "$($NugetRepositoryUrl.TrimEnd('/'))/package/chocolatey.1.2.1.nupkg" # This url should result in an immediate download when you navigate to it # $RequestArguments.Credential = $NugetRepositor圜redential # ("password" | ConvertTo-SecureString -AsPlainText -Force) # If required, add the repository access credential here $NugetRepositoryUrl = "INTERNAL REPO URL" # Should be similar to what you see when you browse Your internal repository url (the main one). # We use this variable for future REST calls. ::SecurityProtocol = ::SecurityProtocol -bor 3072 # installed (.NET 4.5 is an in-place upgrade). NET 4.0, even though they are addressable if. # Use integers because the enumeration value for TLS 1.2 won't exist # Set TLS 1.2 (3072) as that is the minimum required by various up-to-date repositories. # We initialize a few things that are needed by this script - there are no other requirements. # You need to have downloaded the Chocolatey package as well. Download Chocolatey Package and Put on Internal Repository # # repositories and types from one server installation. # are repository servers and will give you the ability to manage multiple # Chocolatey Software recommends Nexus, Artifactory Pro, or ProGet as they # generally really quick to set up and there are quite a few options. # You'll need an internal/private cloud repository you can use. Internal/Private Cloud Repository Set Up # # Here are the requirements necessary to ensure this is successful. ![]() Your use of the packages on this site means you understand they are not supported or guaranteed in any way. With any edition of Chocolatey (including the free open source edition), you can host your own packages and cache or internalize existing community packages. Packages offered here are subject to distribution rights, which means they may need to reach out further to the internet to the official locations to download files at runtime.įortunately, distribution rights do not apply for internal use. If you are an organization using Chocolatey, we want your experience to be fully reliable.ĭue to the nature of this publicly offered repository, reliability cannot be guaranteed. Human moderators who give final review and sign off.Security, consistency, and quality checking.ModerationĮvery version of each package undergoes a rigorous moderation process before it goes live that typically includes: Welcome to the Chocolatey Community Package Repository! The packages found in this section of the site are provided, maintained, and moderated by the community. ![]()
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