QuikChange Multi Site-Directed Mutagenesis Kit

Site Directed Mutagenesis (SDM) Human - Point mutation BxPC-3 uPAR

Experiment
Site Directed Mutagenesis (SDM) Human - Point mutation BxPC-3 uPAR
Product
QuikChange Multi Site-Directed Mutagenesis Kit from Agilent Technologies
Manufacturer
Agilent Technologies

Protocol tips

Upstream tips
- XL10-Gold cells are resistant to tetracycline and chloramphenicol. If the mutagenized plasmid contains only the tetR or camR resistance marker, an alternative strain of competent cells must be used
Protocol tips
- Use only the Dpn I enzyme provided; do not substitute with an enzyme from another source.

Publication protocol

Plasmid constructs
A 450-bp fragment of the uPAR promoter ( −402/+48 region) and the luciferase gene present in the pAd-TrackuPARLuc [3] were excised and cloned into a pGEM-T vector (puPARLuc). The reporter plasmid puPARLuc-NF-κBmut was generated by directed mutagenesis. The NF-kB binding site in the uPAR promoter (located at −45 bp of transcription initiation site) 5'-GGGAGGAGT-3' [13], was mutated to 5'-GGATCCAGT-3', using the QuikChange Multi Site-Directed Mutagenesis Kit (Agilent, CA USA), as recommended by the manufacturer and using the mutagenesis primer: 5'-ctctttcgcaaaacgtctggatccagtccctggggccacaaaac-3'. puPARE1A was generated by cloning the uPARE1A construct in a pGEM-T vector. The NF-κBmutation in the puPARE1A to generate puPARE1A-NF-κBmut was carried out by directed mutagenesis as described above. 3xκB-Luc reporter plasmid was previously described [31].

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Papers

Check out relevant papers found by Labettor's AI that are relevant for performing Site Directed Mutagenesis (SDM) Human - Point mutation BxPC-3 uPAR using QuikChange Multi Site-Directed Mutagenesis Kit from Agilent Technologies.

Paper title
AduPARE1A and gemcitabine combined treatment trigger synergistic antitumor effects in pancreatic cancer through NF-κB mediated uPAR activation
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Manufacturer protocol

Download the product protocol from Agilent Technologies for QuikChange Multi Site-Directed Mutagenesis Kit below.

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