The −1313/+26 human KISS1-pGL2-luciferase plasmid (hKiss-Luc) (Mueller et al., 2011 (link)) was kindly provided by Alejandro Lomniczi and Sergio Ojeda. The −4058/+455 mouse Kiss1-pGL4-luciferase plasmid (Atkin et al., 2013 (link)) was kindly provided by Dr. Steven Kliewer (UT Southwestern). We subcloned the −4058/+455 mouse Kiss1 promoter sequence from the pGL4 backbone into a pGL2 backbone using the KpnI-HF, XhoI, and SalI-HF restriction enzymes in 10X Cut Smart Buffer (New England BioLabs) and religated using Quick Ligase Kit (New England BioLabs). This resulted in the −4058/+455 mouse Kiss1-Luciferase-pGL2 plasmid (mKiss-Luc). The murine −1 kb cFos-pGL3-Luciferase (cFos-Luc) (Ely et al., 2011 (link)) and the cFos-Luc with cis-mutations at −59 (μ−59), −313 (μ−313), and at both −59 and −313 (μ−313x-59) bp were previously described (Hoffmann et al., 2018 (link)). The reporter plasmid containing β-galactosidase constitutively driven by the Herpes virus thymidine kinase promoter (TK-βgal) was used to control for transfection efficiency. We obtained the SIX3-pSG5 expression plasmid from Origene Technologies (Rockville, MD) and the murine cFOS-CMV expression plasmid (cFOS) was previously described (Glidewell-Kenney et al., 2013 (link)). Sequences of all plasmids were confirmed by Sanger Sequencing (Eton Bioscience).
Six3 psg5 expression plasmid
Manufactured by OriGene
The SIX3-pSG5 expression plasmid is a laboratory tool that allows for the expression of the SIX3 gene. The plasmid contains the necessary regulatory elements to facilitate the production of the SIX3 protein in a variety of cell types.
Automatically generated - may contain errors
2 protocols using six3 psg5 expression plasmid
1
Generation and Characterization of Kiss1 and cFos Luciferase Reporters
2
Generation and Characterization of Kiss1 and cFos Luciferase Reporters
Check if the same lab product or an alternative is used in the 5 most similar protocols
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The −1313/+26 human KISS1-pGL2-luciferase plasmid (hKiss-Luc) (Mueller et al., 2011 (link)) was kindly provided by Alejandro Lomniczi and Sergio Ojeda. The −4058/+455 mouse Kiss1-pGL4-luciferase plasmid (Atkin et al., 2013 (link)) was kindly provided by Dr. Steven Kliewer (UT Southwestern). We subcloned the −4058/+455 mouse Kiss1 promoter sequence from the pGL4 backbone into a pGL2 backbone using the KpnI-HF, XhoI, and SalI-HF restriction enzymes in 10X Cut Smart Buffer (New England BioLabs) and religated using Quick Ligase Kit (New England BioLabs). This resulted in the −4058/+455 mouse Kiss1-Luciferase-pGL2 plasmid (mKiss-Luc). The murine −1 kb cFos-pGL3-Luciferase (cFos-Luc) (Ely et al., 2011 (link)) and the cFos-Luc with cis-mutations at −59 (μ−59), −313 (μ−313), and at both −59 and −313 (μ−313x-59) bp were previously described (Hoffmann et al., 2018 (link)). The reporter plasmid containing β-galactosidase constitutively driven by the Herpes virus thymidine kinase promoter (TK-βgal) was used to control for transfection efficiency. We obtained the SIX3-pSG5 expression plasmid from Origene Technologies (Rockville, MD) and the murine cFOS-CMV expression plasmid (cFOS) was previously described (Glidewell-Kenney et al., 2013 (link)). Sequences of all plasmids were confirmed by Sanger Sequencing (Eton Bioscience).
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