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We narrowed to 79 results for: plasmid cloning by pcr

Showing: 31 - 40 of 79 results

  1. DNA Purification Without a Kit

    Type
    Blog Post
    Published
    Feb. 11, 2020, 2:15 p.m.
    ...Read our Plasmids 101 blog series Find other plasmid protocols and tips Browse our plasmid cloning blog posts...alkaline lysis plasmid miniprep Starting material: 2 mL bacterial culture Product: plasmid DNA This kit-...kit-free plasmid miniprep protocol from Addgene follows a similar workflow as a column-based plasmid extraction... or agarose gel slices Product: plasmids or DNA fragments The plasmid purification and DNA gel extraction...purification Starting material: PCR product or agarose gel slices Product: purified PCR product or DNA fragments...from the column, as detected by qPCR, but this did not reduce cloning efficiencies.  When compared to ...depending on the type of DNA you’re purifying (plasmid, genomic, or DNA fragments from agarose).    In...

  2. Tips for CRISPR Gene Editing in Mice

    Type
    Blog Post
    Published
    June 28, 2016, 10:59 a.m.
    ... can clone the PCR products into a pBluescript cloning vector and sequence the resulting plasmid. It ... target plasmid and pX330-sgRNA plasmid (Mashiko et al. 2013). The pCAG-EGxxFP target plasmid contains...pCAG-EGXXFP plasmid along with the individual gRNAs in their PX330 plasmids (or similar plasmids for gRNA...can be placed in multi-cloning site (MCS) between EGFP fragments. The pX330 plasmid contains humanized Cas9...sgRNA can be cloned directionally into the BbsI site. (c) The pCAG-EGxxFP target plasmid was co-transfected...insert this region into a pCAG-EGXXFP plasmid using standard cloning techniques (The XX indicates where ...pCAG-EGXXFP plasmid? They are the perfect primer sets to use to genotype your pups. A simple PCR with these...

  3. Plasmids 101: What is a plasmid?

    Type
    Blog Post
    Published
    April 2, 2020, 6:17 p.m.
    ...plasmids, expression plasmids, gene knock-down plasmids, reporter plasmids, viral plasmids, and genome engineering... of scientists and plasmid lovers - serves as an introduction to plasmids. Plasmids 101 will provide you... chosen based on the plasmid you want to clone into. Regardless, once the cloning steps are complete, ...utilize plasmids across a wide range of applications. Some common plasmid types include cloning plasmids...be asked to design, modify, or construct a plasmid. A plasmid is a small circular piece of DNA found in... components that make up a plasmid and why each is important. Our “Plasmids 101” series designed to educate...advancing the field.   What is a plasmid? At their most basic level, plasmids are small circular pieces of...

  4. Plasmids 101: Blue-white Screening

    Type
    Blog Post
    Published
    June 4, 2015, 1:03 p.m.
    ...inserted it into a plasmid, creating an α-complementation cloning vector. When a cloning reaction goes to...This post is part of our ongoing Plasmids 101 series. Plasmids 101 will provide you with an overview ...of the Plasmids 101 posts here. Now that we have covered antibiotic selection here at Plasmids 101, we...screening your cloning reaction. Being able to select for colonies that contain your plasmid is a great start... start when cloning, but how about being able to choose those that contain plasmid with an insert? Blue-white...Make sure you use a proper plasmid (i.e. contains the α-complementation cloning MCS): pGEM-T, pUC18 and ...successful cloning and not because the cell failed to take up the α-complementation plasmid. This way ...

  5. Tag Your Favorite Yeast Genes with Ease

    Type
    Blog Post
    Published
    Nov. 19, 2013, 2:37 p.m.
    ...Additionally, Dr. Formosa has deposited six plasmids with a multiple cloning site in place of the epitope tag for...protocol is surprisingly simple, requiring only a PCR product containing the modifying sequence flanked...to the chromosomal site of insertion. The linear PCR product is introduced into the cell by direct transformation...Addgene distributes several ready-to-use, modular plasmids, combining fluorescent tags, epitope tags, protease...without the artifacts sometimes associated with plasmid-based overexpression. Tim Formosa, at the University...selection marker (KanMX, HphMX, or His3MX).  Each PCR product from this collection will yield an insert...Jürg Bähler have deposited a large collection of plasmids for genome modification in yeast developed by ...

  6. PCR Applications for Copy Number Variation Assays

    Type
    Blog Post
    Published
    April 26, 2022, 1:15 p.m.
    ...to focus on PCR-based applications (qPCR and ddPCR.) qPCR Quantitative/real-time PCR (qPCR) has been used...Reaction (PCR) Protocol: How to Design a primer  Molecular Biology Reference Molecular Cloning Techniques... courtesy of UPODMG 1516 irivbrro ddPCR  Droplet digital PCR (ddPCR) works by using oil droplets to partition...used for copy number assays for years. qPCR works like regular PCR, just with signaling molecules added ...partition a single sample into thousands of PCR reactions using Taqman chemistry (see image, left, of ... Resources on the Addgene blog   Droplet Digital PCR for AAV QuantificationPolymerase Chain Reaction Overview...Analyzing Copy Number Variation with Droplet Digital PCR. Methods Mol Biol. 2018;1768:143-160. doi: 10.1007...

  7. Fluorescent Tagging of Endogenous Genes with SapTrap

    Type
    Blog Post
    Published
    Oct. 12, 2017, 2:26 p.m.
    ...template (Fig. 1, Step 1). There is no need for PCR or cloning, as digestion of the destination vector with...as a universal donor plasmid. The CRISPaint toolkit includes several donor plasmids for protein tagging...polymerase chain reaction (PCR)-based system that allows users to generate a plasmid containing a direct fusion...and a second plasmid containing the homology arms and an N- or C-terminal tag. The two PCR reactions are...only be detected by labor-intensive processes like PCR or by evaluating visual phenotypes. Improving C. ...The Jorgensen Lab recently developed a modular plasmid assembly toolkit called SapTrap to improve CRISPR...repair template. SapTrap includes a prebuilt donor plasmid library containing several types of fluorescent...

  8. Cloning Mammalian Cells with the Agarose Method

    Type
    Blog Post
    Published
    Sept. 7, 2017, 12:17 p.m.
    ...dried out and died. The alternative to cloning rings, dilution cloning into 96-well plates, is extremely time...give rise to single clones, and thus many plates must be examined for single clones and then handled. Additionally... to the expense of dilution cloning. Taking a page from bacterial cloning, we reasoned that we should...cell clones with sterile truncated pipet tips. Indeed, an experienced operator can pick 48 clones from...Tips for using the agarose cloning method Test your agarose temperature. Cloning cells using an agarose overlay...late-arising clones, and making sure we always maintain a backup replicates of each clone in case of contamination...individual cell clones with different expression characteristics from a population of transfected clones. The technique...

  9. Addgene's Tips for Plasmid Quality Control

    Type
    Blog Post
    Published
    Jan. 14, 2016, 3:30 p.m.
    ...element cloned into the plasmid) and any other important features that differentiate this plasmid from its...sharing plasmids through Addgene is that we provide an added level of confidence in the plasmids we distribute...ensuring plasmid quality is to stay organized. Here at Addgene, we collect data on every plasmid we distribute...restriction sites used during cloning, and most importantly, the predicted full plasmid sequence. Other information...during the cloning, sample handling, or sequence assembly steps. All of our incoming plasmids are sequenced...sequence analysis, plasmid verification, and using NCBI BLAST. Tips for high-throughput plasmid sequencing If...on the Addgene Website View our Plasmid Reference Pages Get Plasmid Backbone Information from Our Vector...

  10. What's the Best Way to Elute and Store Your Plasmid DNA?

    Type
    Blog Post
    Published
    June 4, 2020, 1:22 p.m.
    ...fragmenting plasmid DNA, PCR product, or chromosome. One of the main causes of degradation during plasmid DNA ... a kit Read our blog posts about plasmid cloning Browse our Plasmids 101 blog posts Resources on Addgene.org...Biology Reference Find protocols on plasmid cloning Find plasmids for your research ...Nowadays, the most common E. coli strains used for plasmid amplification are endA negative, meaning the gene...degradation and which experiments you can use your plasmid.  DNA elution choices: TE, Tris buffer, or water...polymerases. In the situation where you need to use plasmid in TE for any enzymatic reaction, an effective ...choices that companies offer for the elution of plasmid DNA through their kit. The advantage of using a...
Showing: 31 - 40 of 79 results