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. Author manuscript; available in PMC: 2017 Jun 8.
Published in final edited form as: Cell Host Microbe. 2016 May 16;19(6):891–900. doi: 10.1016/j.chom.2016.05.004

Figure 2.

Figure 2

Characterization of parental and recombinant ZIKVs in cell culture. (A) Plaque morphology of parental and recombinant ZIKVs. (B & C) Comparison of growth kinetics in Vero and C6/36 cells, respectively. Vero and C3/36 cells were infected with parental and recombinant virus at an MOI of 0.01. Viral titers were measured at indicated time points using plaque assays on Vero cells. Means and standard deviations from three independent replicates are shown. Statistics were performed using unpaired student's t-test. *significant (p value < 0.05); **highly significant (p value < 0.01). L.O.D., limitation of detection (100 PFU/ml). (D) An engineered genetic marker in the recombinant ZIKV. An SphI cleavage site, located in the viral E gene of parental virus, was knocked out in the cDNA clone to serve as a genetic marker to distinguish between recombinant virus and parental virus. A 1250-bp fragment (from nucleotides 1,303 to 2,552) spanning the SphI site was amplified using RT-PCR from RNA extracted from either recombinant virus or parental virus. The RT-PCR fragments were subjected to SphI digestion. The 1250-bp fragment derived from recombinant virus should not be cleavable by SphI; whereas the RT-PCR fragment amplified from parental viral RNA should be cleavable by SphI. The expected sizes of the digestion products are indicated. (E) Agarose gel analysis of SphI digestion products. Expected digestion pattern was observed as depicted in panel (D) was observed. The lengths of DNA fragments are indicated on the right side of the agarose gel.