Poplar Plastid-mediated DsSRP54K Against Plagiodera Versicolora

Poplar(Populus L.)is an important woody plant that is planted in large numbers because of its rapid growth and its ability to meet the need for timber for economic construction.The leaf beetle(Plagiodera versicolora)is an insect of the leaf beetle family of Coleoptera,which is one of the main pests of Populus plants.They can feed on poplar leaves and affect the normal growth and development of poplar trees.A new pest control method,RNA interference(RNAi),is a double-stranded RNA(dsRNA)-mediated silencing of target genes,which can be highly effective against pests.A previous report has screened for the RNAi efficient lethal gene SRP54 K of the leaf beetle,encoding signal recognition particle protein 54 K,which is associated with protein transport.In this paper,we used plastid genetic engineering to express dsSRP54 K in poplar plastidswith high lethality against the leaf beetle,and developed a new strategy of using poplar plastid-mediated RNAi against insects.The research content and results mainly include:1.Using biolistic gun transformation method to obtain plastid transformation material of sansin poplar expressing dsSRP54 K in plastidsThe poplar plastid transformation vector pYY120 was constructed,and two resistant shoots capable of expressing dsSRP54 K were obtained by biolistic transformation method;Southern blot results showed that the two plastid-transformed mountain new poplars had reached homogenization;Northern blot results showed that the two plastid-transformed mountain new poplars were able to express dsSRP54 K and the expression amount accounted for 0.2%-0.4% of the total plant RNA.2.The leaf beetle bioassay by plastid-transformed poplarPlastid-expressing dsSRP54 K mountain new poplar was placed in a plant incubator for 8 weeks,and isolated leaves were taken and fed to the leaf beetle larvae at different ages to record insect resistance test data.The results showed that the leaf beetle larvae had high lethality on day 5 of feeding on plastid-transformed poplar.Young leaves of plastid-expressing dsSRP54 K mountain new poplar had 100%mortality on the 5th day of feeding by first instar larvae of the leaf beetle,and first instar larvae feeding on mature and senescent leaves had 100% mortality on the 6th day.3.The leaf beetle bioassay by nuclear-transformed poplarThe nuclear-transformed poplar vector was constructed,and 26 nucleartransformed strains were obtained by Agrobacterium transformation,and the strains #7and #24 with the highest expression were selected for production by QRT identification.The results showed that the nuclear-transformed poplar had no effect on the control of willow blue leaf beetle.In conclusion,poplar plastidscan express and accumulate dsSRP54 K,and poplar plastid-mediated RNAi has good anti-insect effects on the leaf beetle larvae larvae at different ages,while nuclear transfer poplar has no control effect,proving that poplar plastid-mediated dsSRP54 K is an efficient and safe new anti-insect strategy to control the forestry pest the leaf beetle.