Schneider PN, Slusarski DC, Houston DW. (2012) Differential role of axin RGS domain function in Wnt signaling during anteroposterior patterning and maternal axis formation. PLoS One, 7(9):e44096.
Axin is part of a multi-protein complex that facilitates the phosphorylation and degradation of β-catenin, a critical transcription regulator involved in development. In the presence of Wnt signaling, axin is removed from its role in the β-catenin destruction complex, and β-catenin is able to enter the nucleus to activate gene transcription. In this paper, the authors investigate an RGS (Regulator of G-protein Signaling) domain within axin to see if it has a larger role in regulating activity of the β-catenin destruction complex. The authors do this by mutating the RGS domain of axin, while leaving the other adenomatous polyposis coli (APC, a protein that helps regulate how cells divide) interactions and degradation functions intact. The mutant is studied in vitro
and in vivo
using Xenopus laevis
and Danio rerio
. To confirm that the axin mutant is still able to interact with APC and the destruction complex, the authors cloned a 459 bp portion of the Xenopus
APC producing the axin-interacting region. The authors were able to easily generate the control Xenopus
APC fragment by ordering it as a single gBlocks® Gene Fragment
and cloning it into a plasmid containing an HA tag sequence. The expressed fragment was then used to confirm that the wild type and mutant axin RGS domains were able to interact normally with APC.
gBlocks® Gene Fragments are double-stranded, sequence-verified, DNA genomic blocks, 125–3000 bp in length, that can be shipped in 2–5 working days for affordable and easy gene construction or modification. gBlocks Gene Fragments have been used in a wide range of applications including CRISPR-mediated genome editing, antibody research, codon optimization, mutagenesis, and aptamer expression. They can also be used for generating qPCR standards. With gBlocks Gene Fragments, IDT provides a tool that accelerates your research and makes synthetic biology easier and more accessible than ever, for any lab.
Learn more about gBlocks Gene Fragments at www.idtdna.com/gblocks.
Read how gBlocks Gene Fragments have been applied in other research projects:
Towards next generation biosensors—See how fluorescent protein-based Ca2+ indicators based on naturally occurring substrates are assembled using gBlocks Gene Fragments. These sensors are being developed to monitor in vivo neural activity.
Site-directed mutagenesis—improvements to established methods—Site-directed mutagenesis techniques have relied primarily on PCR and standard cloning methods. Read about some of the common cloning methods used for mutagenesis and how double-stranded DNA fragments (gBlocks Gene Fragments) can save you both time and money.
Easily-Designed Standard Curves for qPCR—Use this easy way to combine control templates/multiple targets onto a single construct, and the advantages that it can provide for PCR experiments.
Author: Ellen Prediger, PhD, is a senior scientific writer at IDT.
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