Growth Characteristics Of Inhibition Of Expression Of BpGLK Betula Pendula ‘Dalecarlica’ And Preliminary Evaluation Of Environmental Safety

Inhibition of expression of Bp GLK Betula pendula ‘Dalecarlica’ with bright yellow-green leaf color in summer,which has a promising development in urban greening applications.As a prerequisite for the application and promotion of transgenic forest trees,it is not only necessary to understand the impact of the introduction of exogenous genes on their growth and wood properties,but also to detect the impact of exogenous genes on soil and soil microorganisms,and to evaluate the environmental safety of transgenic forest trees.Therefore,this experiment was conducted to analyze the growth,leaf color and wood properties of the transgenic strains by using the Bp GLK inhibition expression birch,and determine soil physicochemical properties,soil enzyme activity,soil microbial quantity,and rhizosphere soil microbial community structure.At the same time,detect whether the imported exogenous gene(herbicide resistant Bar gene)has undergone horizontal transfer,The research results lay a theoretical foundation for the promotion and application of Bp GLK inhibiting the expression of Betula pendula ‘Dalecarlica’ in landscaping in the later stage.The specific results are as follows:(1)Using 2-year-old and 3-year-old Bp GLK inhibited expression birch(RE1-RE8)as materials,PCR and q RT-PCR analysis showed that the introduced Bp GLK interference sequence and Bar gene were still integrated into the genome of the transgenic strains,and both showed downregulation of Bp GLK gene expression at m RNA expression levels.(2)The investigation on leaf color and photosynthetic pigment of Bp GLK inhibiting the expression of Betula pendula ‘Dalecarlica’ was carried out for two consecutive years.The L*and b* values of RE1-5 strains were significantly higher than RE6-8 strains and WT strains.The determination of photosynthetic pigments showed that the chlorophyll a,chlorophyll b,and total chlorophyll contents of the RE1-RE5 strains were lower than those of the RE6-RE8 and WT strains.Analysis using q RT-PCR technology showed that the genes involved in photosynthesis,and the genes related to chlorophyll synthesis,were significantly downregulated in the leaves of yellow-green leaf strains.(3)The investigation on height growth of 2-year-old Bp GLK inhibited expression of Betula pendula ‘Dalecarlica’ showed that only RE4 and RE6 strains were significantly higher than WT strains,and other strains were not significantly different from WT strains.The results of wood property analysis showed that in terms of wood density,fiber length width ratio,hemicellulose and lignin content,there was no significant difference between most RE strains and WT strains,but in terms of cellulose content,five RE strains(RE5,RE3,RE2,RE8,RE7)were significantly lower than WT strains.(4)Using Bp GLK to inhibit the expression of Betula pendula ‘Dalecarlica’ soil as the material,the physicochemical properties of RE strains showed that the soil p H value,organic carbon,total nitrogen,and available nitrogen content were lower or significantly lower than those of WT strains.The contents of available phosphorus,total potassium,and available potassium were significantly higher in the WT strains(except for RE1).The results of soil enzyme activity measurement showed that urease and sucrase reached their highest levels in mid July,and neutral protease was the lowest in mid July.In mid September,the soil urease and sucrase activities of the RE strains were lower or significantly lower than those of the WT strains,while neutral protease activity was significantly higher than that of the WT strains.The results of the determination of the number of bacteria,fungi,and actinomycetes in the soil showed that the number of soil microorganisms in 3-year-old Betula pendula ‘Dalecarlica’ was higher than that in 2-year-old Betula pendula ‘Dalecarlica’.And there are seasonal changes in the number of microorganisms.(5)The rhizosphere soil of Betula pendula ‘Dalecarlica’(RE1-3 strains)with Bp GLK inhibiting expression was used as materials,analyzing the structure of bacterial and fungal communities through high-throughput sequencing technology.The results showed that the bacterial community structure of the transgenic strains was less different from that of the WT strains,and the fungal community structure of the transgenic strain was more different from that of the WT strains.The above results indicate that the dominant bacterial genera in the rhizosphere soil of RE strains are not entirely the same during different growth periods,but all dominant bacterial genera are beneficial bacteria,and no adverse effects on the growth and development of RE strains have been found.(6)Using the DNA of weeds and soil microorganisms around the RE strains as a template,PCR amplification results showed that the Bp GLK interference sequence and Bar band were not amplified in the genomes of weeds and soil microorganisms,indicating that the above genes did not undergo horizontal transfer.In conclusion,at the age of 2 and 3 years,the interference fragment of Bp GLK and the resistance Bar gene of Betula pendula ‘Dalecarlica’ schizophylla are still integrated into its genome.The leaves of the previously obtained RE1～RE5 strains are still bright chartreuse in the growing season.Most transgenic strains have not affected their growth and wood properties due to the low expression of Bp GLK gene.The physical and chemical properties of soil,soil enzyme activities,and the number of soil microorganisms.Although the microbial community structure has had a certain impact,no adverse effects have yet been identified.