Basic Amino Acid Mutations in the Nuclear Localization Signal of Hibiscus Chlorotic Ringspot Virus p23 Inhibit Virus Long Distance Movement
Figure 5
Less severe symptoms were observed in the newly emerged leaves from amiRp23-inoculated kenaf plants which were pre-infected with HCRSV 10 days earlier.
(A) Overlapping PCR to obtain amiRp23 or amiRSO. The a, b, c represent the required fragments to form amiRp23 or amiRSO. PCR fragment d is the product of overlapping PCR from a, b and c. (B) Verification of amiRp23 and amiRSO transient expression in the kenaf leaves using Agrobacterium-infiltration by qRT-PCR. Mock, amiRp23 and amiRSO represent total RNA from mock, amiRp23 and amiRSO Agro-infiltrated kenaf leaves, respectively. Verification of amiRp23 (C) and p23 gene (D) transcript levels in the HCRSV-infected kenaf leaves treated with mock, amiRp23 and amiRSO by qRT-PCR, respectively. V+mock, V+amiRp23 and V+amiRSO represent total RNA from HCRSV infected leaves followed by inoculation with mock, amiRp23 and amiRSO, respectively. (E) Comparison of symptoms (red circles with dotted outline) in the newly emerged leaves between mock (more severe) and amiRp23-inoculated kenaf leaves (less severe) pre-inoculated with HCRSV 10 days earlier, using visual observations. Significant differences of relative amiRp23 and p23 gene between mock with amiRp23 and mock with amiRSO treated plants were calculated using the one sample Student’s t-test. Asterisks (∗∗) indicate significance at 0.01 levels of confidence.