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Figure 1
Overexpression, purification and identification using a Western blot of recombinant C1ORF123 protein. (a) Analysis of the overexpressed and Ni2+–NTA affinity-purified rC1ORF123 protein using SDS–PAGE (12%). Lane 1, PageRuler prestained protein ladder (Thermo Scientific, USA; labelled in kDa); lanes 2, 3 and 4, total protein, pellet and supernatant of crude extract before IPTG induction, respectively, as a negative control; lanes 5, 6 and 7, total protein, pellet and supernatant of crude extract after IPTG induction, respectively; lane 8, rC1ORF123 purified using Ni2+–NTA affinity chromatography. (b) Size-exclusion chromatography (SEC) results using a HiLoad 16/600 Superdex 75 pg gel-filtration column (GE Healthcare, UK). The rC1ORF123 protein eluted as a single peak between those for myoglobin (17 kDa) and ovalbumin (44 kDa), suggesting that the rC1ORF123 protein may be a monomer in solution. rC1ORF123 purified by SEC is shown on 12% SDS–PAGE, with the protein band corresponding to ∼20 kDa. (c) Calibration curve for size-exclusion chromatography on a Superdex 16/600 75 pg gel-filtration column indicating the molecular weight of rC1ORF123 to be ∼28 kDa. (d) The recombinant rC1ORF123 protein was positively detected by a Western blot using anti-human C1ORF123 antibodies (right panel); the band at ∼20 kDa corresponding to purified rC1ORF123 is shown (left panel).

ISSN: 2053-230X
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