The protein domain microarray was conducted by the Protein Array and Analysis Core (PAAC) at MD Anderson Cancer Center. A comprehensive library of human protein domains that potentially read methyllysine marks was cloned into a pGEX vector by Biomatik (Cambridge, Canada) using gene synthesis to best optimize the open reading frames for bacterial expression. Escherichia coli was used to express the protein domains as GST fusions, which were purified using glutathione-Sepharose beads. The recombinant domains were arrayed onto nitrocellulose-coated glass slides (Oncyte®Avid slides, Grace Bio-Labs, Bend, OR), using an Aushon 2470 pin microarrayer, as previously described (Espejo et al., 2002 (link)). Biotinylated compounds were pre- conjugated to streptavidin-Cy3 to generate fluorescent probes. Following probing, fluorescent signal was detected with a GenePix 4200A Microarray Scanner (Molecular Devices).
2470 pin microarrayer
Manufactured by Grace Bio-Labs
The 2470 pin microarrayer is a laboratory instrument used for the precise and automated spotting of small liquid samples onto a solid surface, such as a microarray slide. The device utilizes a series of pins to transfer and deposit the samples in a high-density array format. The core function of the 2470 pin microarrayer is to facilitate the creation of microarrays for applications in areas such as genomics, proteomics, and drug discovery.
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2 protocols using 2470 pin microarrayer
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Protein Domain Microarray Analysis
2
Protein Domain Microarray Analysis
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The protein domain microarray was conducted by the Protein Array and Analysis Core (PAAC) at MD Anderson Cancer Center. A comprehensive library of human protein domains that potentially read methyllysine marks was cloned into a pGEX vector by Biomatik (Cambridge, Canada) using gene synthesis to best optimize the open reading frames for bacterial expression. Escherichia coli was used to express the protein domains as GST fusions, which were purified using glutathione-Sepharose beads. The recombinant domains were arrayed onto nitrocellulose-coated glass slides (Oncyte®Avid slides, Grace Bio-Labs, Bend, OR), using an Aushon 2470 pin microarrayer, as previously described (Espejo et al., 2002 (link)). Biotinylated compounds were pre- conjugated to streptavidin-Cy3 to generate fluorescent probes. Following probing, fluorescent signal was detected with a GenePix 4200A Microarray Scanner (Molecular Devices).
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