Extraction and bioactivity analysis of total protein of K326 tobacco leaves with high expression of Nicotiala alata defensin 1 gene / 中国生物制品学杂志

ABSTRACT

@#Objective To extract the total protein of K326 tobacco leaves with high expression of Nicotiana alata defensin 1(NaD1) gene and analyze its bioactivity.Methods Total proteins were extracted from Nicotiana alata flowers,wild type(WT) and K326 tobacco leaves(transgenic) with high expression of NaD1 gene,and determined for the concentrations by Bardford method,while for the antibacterial activity against fungi by filter paper method,and for the inhibition activity on cancer cells(HeLa cells) by CCK-8.Results The total protein concentrations of Nicotiana alata flowers,WT and transgenic K326 tobacco leaves were 11.25,10.33 and 10.14 mg/mL,respectively.The antibacterial activity of total protein from transgenic K326 tobacco leaves against Candida albicans was(85.68±3.08)%,which was 1.33 and 1.14 times that of total protein from WT K326 tobacco leaves and Nicotiana alata flowers,respectively(F=15 339,P <0.05);The antibacterial activity against Fusarium oxysporum was(148.48±2.47)%,which was 1.09 and 1.08 times that of total protein from WT K326tobacco leaves and Nicotiana alata flowers,respectively(F=4.927,P <0.05).The IC_(50) value of transgenic K326 tobacco leaf protein on HeLa cells was the smallest(6.11 mg/mL),and the inhibitory activity was 1.56 and 1.21 times that of total protein of WT K326 tobacco leaves and Nicotiana alata flowers,respectively(F=89.748,P <0.05).Conclusion The total protein of K326 tobacco with high expression of NaD1 gene has good antibacterial and anticancer bioactivities,which provides an experimental basis for producing antibacterial and anticancer biological agents with tobacco as bioreactor.