Open Access
Research Article

Figure 1.

image

Download original image

Selection and identification of the stably expressed cell line. (A) Schematic diagram of vector construction. Replacement of the GFP with a sequence containing 5′ IgK SP and 3′ myc-His sequence using an isocaudamer technique resulted in the creation of pBobi-IMHIP vector. Lentiviral vector pBobi-cathB1 was generated by insertion of Ac-cathB-1 coding sequence excluding SP into the pBobi-IMHIP vector by XbaI and BamHI restriction sites. (B) Lentiviral packaging of pBobi-cathB1 and pBobi-GFP. Under the fluorescence microscope, more than 90% of cells in the pBobi-GFP transfected group displayed green fluorescence. (C) Assessment of the establishment of 293T-cathB1. Anti-Myc IF staining was performed in the two cell lines. All cells in 293T-cathB1 showed red fluorescence representing 100% positive, while all cells in 293T-GFP showed no fluorescence under the red fluorescent filter due to the lack of myc-tag expression. (D) Tests for rAc-cathB-1 expression. Total RNA and protein samples were extracted from two groups of cells. Results of RT-PCR (top two panels) and western blot (lower two panels) showed that rAc-cathB-1 was highly expressed in 293T-cathB1 at both mRNA and protein levels. Actb, RT-PCR control and β-actin, loading control for western blot; WL, white light; and bar = 100 μm.

Current usage metrics show cumulative count of Article Views (full-text article views including HTML views, PDF and ePub downloads, according to the available data) and Abstracts Views on Vision4Press platform.

Data correspond to usage on the plateform after 2015. The current usage metrics is available 48-96 hours after online publication and is updated daily on week days.

Initial download of the metrics may take a while.