The Identification And Functional Confirmation Of Nicotianamine Synthase Gene Horizontal Transferred In Bemisia Tabacci (Hemiptera:Aleyrodidae)

Bemisia tabaci(Hemiptera: aleyrodidae)is a worldwide invasive pest that causes significant losses to agricultural production because of its wide spread,serious harm,difficulty in control.For a deeper understanding of the physiological activities and metabolic processes of whitefly and accomplishing better control of whitefly,we screened genes that may have the function of nicotianamine synthase like BtNAS with the help of previous genome sequencing analysis,In view of its important role and research value in plants,we studied the its function in whitefly.1.BtNAS sequence analysis and determination of Horizontal Transfer EventsDNA exchange through Horizontal Gene Transfer(HGT)is an important driving force in the evolution of multiple species,as it allows receptors to quickly acquire a trait(detoxification...)or adapt to a new environment(improved feeding efficiency).By transcriptome genome alignment and gene cloning,homologous sequence evolution tree analysis,we identified that BtNAS came from plants and was embedded into the whitefly genome through some mechanism.2.BtNAS expression profile analysisWe measured the expression of BtNAS at different developmental ages(nymphs,adults),different parts(head,thoax and abdomen),when treated with different ions(iron,manganese,copper,zinc)and different concentrations of zinc ions in whitefly.The results show that BtNAS is highly expressed in adults and abdomen in whitefly.and can correspond to zinc ions in food.One day treatment with gradient zinc(0,0.2,0.4,0.8,1.0,1.5,2.0,4.0g/L)show that BtNAS is response to the concentration of zinc ion.When the concentration is above 1.0g/L,the expression level of BtNAS is up-regulated.It was found that the expression level of BtNAS was decreased to 70% when treated without zinc ion,and up to 1.5 times with the high zinc ion concentration(2g/L).To further verify the reliability of the results,we found that the results of zinc ion treatment from 1 day to 3 day were consistent with the previous results,indicating that BtNAS was induced by high concentration of zinc ion3 Prokaryotic expression of BtNAS and functional study in vitroThrough the prokaryotic expression in vitro and solubilization of BtNAS with the determination of enzyme activity by liquid-mass spectrometry-mediated,we found that BtNAS can synthesize nicotianamine in vitro,thus confirm that the function of BtNAS is similar to plant and bacteria.4 RNA interference(RNAi)and BtNAS functional research in vivoWe found a significant down-regulated of BtNAS after RNAi treatment and a decrease in the amount of zinc and cobalt under four-days continuous interference treatment.With the combination of RNAi and bioassay,we found that down-regulated the expression of BtNAS could result in the increase of whitefly mortality when treated with excess zinc ion.In combination with the above experimental results,we concluded that BtNAS is involved in the accumulation of zinc and cobalt elements in the body of whitefly and may be involved in the process of super-tolerance of zinc ions.