Cloning and Function of MeABF1 in Cassava

Objective The gene of transcription factor ABF1 from Manihot esculenta Crantz (MeABF1) was cloned, and its prokaryotic expression and disease resistance-related function analyzed.

Method Coding sequence (CDS) of MeABF1 was amplified from the M. esculenta cultivar, South China 124 (SC124). MeABF1-pET32a fusion expression vector was constructed and transformed into DE3 competent cells for protein induction. Expression of the gene was determined by SDS-PAGE and western blotting. Cassava leaves were sprayed with a solution of purified MeABF1 protein and tested for burst of reactive oxygen species (ROS), expression of pathogenesis-related gene (PR), and disease resistance to Xanthomonas axonopodis pv. manihotis (Xam).

Result The CDS sequence of MeABF1 was 1,491bp long. The MeABF1-Myc fusion protein with increasing band intensity over time was detected after 1mmol·L⁻¹ isopropyl β-D-thiogalactoside (IPTG) induction at 37℃ for 1h. The peak ROS burst in the treated leaves was 3-fold of control, and the MePR1 and MePR2 expressions upregulated 3.5-fold and 1.75-fold, respectively. A 2d treatment significantly enhanced the disease resistance to Xam of the cassava leaves. The bacterial counts and lesion area on the leaves 6d post inoculation (dpi) were significantly reduced with improved phenotypic outcomes over control.

Conclusion MeABF1-pET32a was successfully expressed in cassava leaves with the induction by 1mmol·L⁻¹ IPTG at 37°C. The foliar MeABF1 application significantly enhanced the resistance of the leaves to Xam infection with burst of ROS and heightened PR expression.