| Karelinia caspia is a perennial herb of the genus Karelinia in Asteraceae,commonly known as the fat girl and the fat baby grass.It is a desert plant widely distributed in arid and semi-arid areas.Living in desert environment for a long time,Karelinia caspia has strong drought tolerance,high temperature tolerance,salt tolerance and other broad-spectrum stress resistance,so it is an important plant resource for studying plant stress resistance.In order to explore the relationship between genes related to hormone synthesis and high temperature tolerance in Karelinia caspia,we analyzed the effects of habitat,temperature and exogenous auxin on floral organ development in Karelinia caspia.The plant growth hormone biosynthesis differentially expressed gene Kc APSK was screened from the transcriptome sequencing results of Karelinia caspia after high temperature stress in the early stage of our research group,and the full length of Kc APSK gene CDS was cloned.The protein results and expression patterns of Kc APSK gene were analyzed,and the plant expression vector was constructed by Gateway technology.The p K2GW7-Kc APSK expression vector was transformed into tobacco by Agrobacterium-mediated transformation,and a large number of callus were obtained.The results of this paper are as follows:(1)High temperature is beneficial to the development of floral organs in Karelinia caspia,while extreme temperature is detrimental to the development of floral organs.Exogenous auxin IAA is beneficial to flower organ development under extreme high temperature.By measuring the changes in the contents of endogenous hormones in flower organs of Karelinia caspia under different concentrations of auxin(IAA)and female flower styles,male flower filamentes and flower organs under the four habitats of artificial green space,artificial green space,high temperature and high temperature desert at room temperature.The results showed that 27°C?30°C was beneficial to the growth of floral organs in Karelinia caspia during the bud stage,while 38°C?40°C was beneficial to the growth of floral organs in Karelinia caspia during flowering.Applying a certain concentration of IAA promoted the development of floral organs of Karelinia caspia.In desert environment,0.3 ?mol/L IAA promoted the development of floral organs of Karelinia caspia most obviously,while in artificial green land,0.1 ?mol/L IAA promoted the development of floral organs of Karelinia caspia best.Through the above experiments,it was found that plants can alleviate the harm caused by high temperature stress by regulating the content of hormones in their bodies.(2)The expression pattern of plant hormone biosynthesis gene APSK in Karelinia caspia was positively correlated with high temperature stress.According to transcriptome sequencing analysis of Karelinia caspia under high temperature stress,it was found that the expression of endogenous growth hormone biosynthesis related gene APSK was significantly up-regulated.Therefore,the CDS sequence of this gene was cloned in this study,and the sequencing showed that its full length was 870 bp,encoding 290 amino acids,and it was named Kc APSK.The expression pattern of this gene under high temperature stress was analyzed by RT-PCR,the results showed that the expression level of the gene increased first and then decreased with the extension of heat stress time.When the stress time was 8 h,the expression level reached the maximum,and then decreased again from 8 h to 16 h,but it was still higher than the control level.Our results suggest that high temperature can promote the expression of this gene in Karelinia caspia,so it is speculated that this gene may be involved in the positive regulation of Karelinia caspia response to high temperature stress.(3)Kc APSK plant expression vector was constructed and a large number of transgenic tobacco callus were obtained.The obtained target gene Kc APSK was constructed into the plant expression vector p K2GW7 by Gateway technology,and the plant expression vector p K2GW7-Kc APSK was obtained.In addition,the vector obtained was introduced into Agrobacterium tumefacien LBA4404 strain and transformed into tobacco by Agrobacterium tumefacien mediated leaf plate method.A large number of callus were obtained,which laid a foundation for further exploring the function of Kc APSK gene. |
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