Screen And Analysis Of High Glandular Density K326 Material

Tobacco trichomes contribute greatly to leaf quality and plant resistance to biotic and abiotic stress.the NtCycB2,a negative regulate gene of trichome development,this gene knockout increases the density of secretory glandular density.In order to create multi-gland hair tobacco materials,breeding new varieties of directional improvement.In this study,two NtCycB2 knockout strains of K326 and the T1 generation of wu-16 were detected and screened,Obtained homozygous de-carrier mutant.On this basis,a comparative analysis of agronomic traits,glandular hair morphology,chemical composition of leaves,chemical constituents of roasted tobacco and neutral aroma components of two strains were carried out.Laid the foundation for the application of this gene in tobacco breeding.The following results were obtained.1.Detection and analysis of the NtCycB2 knockout positive strains Wu-2 and Wu-16T1,Sequence analysis showed that NtCycB2 in Wu-2 showed two types of mutation:one is 4 bp deletion coupled with a C/T substitution,another is 7 bp deletion;while in Wu-16,only one type of mutation was found:a A/T base-pair insertion.And homozygous knockout plants without transgenic elements were obtained by PCR detection of hygromycin.2.Agronomic traits were compared between K326 Wu-2 and Wu-16,the results showed that:Compared with the K326,Wu-2 and Wu-16 show significant increase in plant height and leaf distance,and the stem circumference,maximum leaf length,maximum leaf width and number of leaves were also slightly increased.3.The glandular trichome morphology and density of upper,middle and lower leaves of K326,Wu-2 and Wu-16 were compared,the results showed that:trichomes on the leaf surface of K326,Wu-2 and Wu-16 were all composed by long-stalk glandular trichomes,short-stalk glandular trichomes and non-secretory trichomes.More interestingly,branched glandular trichomes were observed on the upper leaves of Wu-2.All three materials showed a higher density of the upper leaf gland than the middle leaf was larger than the lower leaf.The glandular hair density of any of the same parts Wu-16 and Wu-2 was higher than that of K326.4.Leaf chemical composition were analyzed by GC-MS,the results showed that K326,Wu-2 and Wu-16 all contain cembranoid,sucrose esters and alkanes.The total amount of Wu-2 and Wu-16 leaf exudates was significantly higher than that of the control K326.Among them,the content of cyanostanes and alkanes was more prominent,while the content of sucrose esters was slightly reduced.5.Comparison of conventional chemical composition of roasted tobacco between K326,Wu-2 and Wu-16 showed that:the contents of Reducing sugars,total sugars,Total nitrogen and Nicotine were all increased in Wu-2 and Wu-16 compared with K326;on the other hand,the content of Potassium and Chlorine in Wu-2 and Wu-16 were lower than that of K326.Neutral aroma component in K326,Wu-2 and Wu-16 were analyzed by GC-MS,and the results showed that the contents of Phenylalanine,Browning products,Chlorphenoids,Neophytadiene and other compontents were all increased in Wu-2 and Wu-16 compared with K326.