Journal: Protein expression and purification

Article Title: Rapid and Efficient Purification of Drosophila Homeodomain Transcription Factors for Biophysical Characterization

doi: 10.1016/j.pep.2019.02.001

Figure Lengend Snippet: Vector map (A) and sequence map (B) of HisSUMO-HD constructs. Each of the HDs for FTZ, ABD-A, ABD-B, ANTP, and UBX were cloned into a T7-inducible HisSUMO vector between KpnI and XbaI restriction sites. (C) Sequence alignment of overexpressed D. melanogaster HDs. The 107 member homeodomain family was submitted to ClustalW [48] using default values and analysed using Jalview [49]. Only the five HDs studied in this paper are shown. Highly conserved residues within similar physicochemical properties in all 107 HDs analysed are shown (red – positive, blue – negative, yellow – polar, green – hydrophobic). Absolutely conserved residues and residues with greater than 70% similarity are labelled with a star or cross, respectively.

Article Snippet: The HD open reading frames of FTZ (Uniprot: {"type":"entrez-protein","attrs":{"text":"P02835","term_id":"25453437","term_text":"P02835"}} P02835 , residues 254–313) and ANTP ( {"type":"entrez-protein","attrs":{"text":"P02833","term_id":"123317","term_text":"P02833"}} P02833 , residues 297–356) were Escherichia coli ( E. coli ) codon-optimized and synthesized by Synbio Technologies and cloned into a modified T7-inducible HisSUMO vector in between compatible Kpn I and Xba I restriction sites ( ).

Techniques: Plasmid Preparation, Sequencing, Construct, Clone Assay

Journal: Protein expression and purification

Article Title: Rapid and Efficient Purification of Drosophila Homeodomain Transcription Factors for Biophysical Characterization

doi: 10.1016/j.pep.2019.02.001

Figure Lengend Snippet: (A) Purification gel of ANTPHD showing samples taken at each step of the purification process (as described in the Materials & Methods). (B) Gel of purified D. melanogaster HDs for FTZ, ABD-A, ABD-B, ANTP, and UBX. Each lane was loaded with 5 ug of purified HD protein. Spectra Multicolor Broad Range Protein Ladder (Thermo-Fisher) was used in both gels to monitor migration and determine molecular weights of the HisSUMO-HD, HisSUMO tag, and isolated HD proteins. Gels were stained with Coomassie Brilliant Blue G-250 and visualized using an iBright FL1000 gel imager.

Article Snippet: The HD open reading frames of FTZ (Uniprot: {"type":"entrez-protein","attrs":{"text":"P02835","term_id":"25453437","term_text":"P02835"}} P02835 , residues 254–313) and ANTP ( {"type":"entrez-protein","attrs":{"text":"P02833","term_id":"123317","term_text":"P02833"}} P02833 , residues 297–356) were Escherichia coli ( E. coli ) codon-optimized and synthesized by Synbio Technologies and cloned into a modified T7-inducible HisSUMO vector in between compatible Kpn I and Xba I restriction sites ( ).

Techniques: Purification, Migration, Isolation, Staining

Journal: Protein expression and purification

Article Title: Rapid and Efficient Purification of Drosophila Homeodomain Transcription Factors for Biophysical Characterization

doi: 10.1016/j.pep.2019.02.001

Figure Lengend Snippet: EMSA of isolated FTZ, ANTP, and ABD- A HDs binding to the consensus sequence 5’-GCGTTTAATTAGCCC-3’ (A-C) and the null sequence 5’-GCGCGCCGGCGGCCC-3’ (D-F).

Article Snippet: The HD open reading frames of FTZ (Uniprot: {"type":"entrez-protein","attrs":{"text":"P02835","term_id":"25453437","term_text":"P02835"}} P02835 , residues 254–313) and ANTP ( {"type":"entrez-protein","attrs":{"text":"P02833","term_id":"123317","term_text":"P02833"}} P02833 , residues 297–356) were Escherichia coli ( E. coli ) codon-optimized and synthesized by Synbio Technologies and cloned into a modified T7-inducible HisSUMO vector in between compatible Kpn I and Xba I restriction sites ( ).

Techniques: Isolation, Binding Assay, Sequencing