| Karelinia caspica is a typical perennial herb,it is belonging to Compositae,Which which is flowering and pollinating from July to August of each year.It is a kind of high temperature tolerant plant with high research value.It is a broad-spectrum resistant plant with high research value.In order to clarify the relationship between the synthesis of organ hormones and its high temperature tolerance,the morphological characteristics of the flower organs of Karelinia caspica in different environments,different temperatures and different concentrations of IAA and GA were determined.The flowers under different environmental conditions were determined.The content of IAA and ABA in flower organs and the contents of endogenous SA,JA and ABA in different concentrations of IAA and GA were studied.The expression patterns of different hormone-related heat-tolerant genes in Karelinia caspica under high temperature stress were analyzed by q RT-PCR.The expression pattern,and the expression patterns of IAA and ABA synthesis related genes in the different days of the flowering organs of Karelinia caspica on the 5th day before flowering and on the day of flowering,and bioinformatics analysis of the differential gene Kcb HLH71 related to auxin hormone synthesis,cloning,expression pattern analysis and construction of overexpression vectors,resulting in the following conclusions:1.Hormones may be involved in the regulation of Karelinia caspica in response to high temperature stress.The diameters of the flowering organs of Karelinia caspica under the four conditions of artificial green space(27~30°C),desert normal temperature(30~33°C),artificial green space high temperature(38~40°C),and desert high temperature(40°C or above)were measured.Stamen and pistil length and pollen activity,it was found that flower organ size,pistil and stamen length and pollen activity decreased with increasing temperature,and stamens were more susceptible to high temperature than pistil,the artificial green space is twice as long as the stamen in the desert at normal temperature and artificial green,and is 1.1 times longer than the stamen in the desert at high temperature.High temperature has an inhibitory effect on the size of stamen flower organs and stamen growth,but it has a promoting effect on the growth of pistil,The artificial temperature of normal temperature artificial soil and normal temperature desert,artificial green space high temperature and desert high temperature of the pistil of Karelinia caspica from on the 5th day before flowering and on the 3th day after flowering,respectively,1.1 times longer,shorter 17% and 25%.The experimental results of different concentrations of hormones showed that when 0.1?M IAA and 60?M GA3 were applied,the organ development of Karelinia caspica was significantly promoted.It is speculated that a certain high temperature(40 ~ 45°C)is conducive to the development of the pistil of the flowering organs of Karelinia caspica,so that the stigma is exposed,which may be related to cross-pollination;in addition,maintaining high levels of auxin hormones under high temperature conditions is conducive to enhancing high temperature tolerance of Karelinia caspica.2.The content of endogenous hormones IAA and ABA in flower organs was determined by LC-MS.The content of endogenous hormones IAA and ABA in floral organs determined by LC-MS showed that: the content of auxin on the day of flowering organs of Karelinia caspica in normal temperature of artificial green space was 20% lower than that in high temperature of artificial green space,and 4 times higher than that in high temperature of desert.The content of abscisic acid on the day of flowering organs of Karelinia caspica in normal temperature of artificial green space was 3% lower than that in high temperature of artificial green space,and 54% lower than that in high temperature of desert.In order to further study how hormones play a role in adversity environment,the contents of SA,JA and ABA in flower organs were measured by LC-MS after applying IAA and GA.It was found that:(1)normal temperature of artificial green space conditions,flowers The contents of ABA and JA in the organs increased firstly and then decreased with the increase of the above-mentioned hormone concentrations,but the overall changes were not significant.The content of SA in the flower organs decreased with the increase of spraying concentration after IAA and GA application.(2)Under desert conditions,the content of ABA in flower organs increased with the increase of the above-mentioned hormone concentrations.The content of JA in flower organs decreased with the increase of concentration after IAA and GA application.The content of SA in the floral organs of the hormones decreased with the increase of the spray concentration.3.The transcriptome analysis under high temperature stress showed that 359 differentially expressed genes in hormone signaling pathway were annotated in leaf organs of Karelinia caspica after high temperature treatment.At the same time,the expression patterns of 10 genes related to different hormone synthesis under high temperature stress were analyzed by q RT-PCR.The expression patterns of 6 genes related to IAA and ABA synthesis at different temperatures were also analyzed on the fifth day before flowering and on the flowering day.The results showed that 12 genes related to hormone synthesis were all related to response to high temperature stress.However,its function still needs further verification in order to provide theoretical support for further research.4.According to the results of transcriptome sequencing analysis of Karelinia caspica under high temperature stress,the growth hormone synthesis related gene b HLH was up-regulated by 10 times compared with the control.The gene was cloned by PCR and named Kcb HLH71.The expression pattern of the gene under high temperature stress was analyzed by q RT-PCR.The results of expression analysis showed that under high temperature stress,the expression of Kcb HLH71 gene was inhibited at 2h,and then increased gradually,reaching a peak at 12 h,which was 2.1 times higher than that of the control.The expression of Kcb HLH71 gene decreased again at 12 h to 24 h,but still higher than that of the control.The above results indicated that high temperature could significantly induce the expression of Kcb HLH71 gene,suggesting that this gene might be involved in the positive regulation of response to high temperature stress in Karelinia caspica.The overexpression vector OE35S-Kcb HLH71 of Kcb HLH71 gene was constructed by seamless cloning technology,which laid a foundation for further verification of the function of Kcb HLH71 gene.In summary: this study found that hormones play an important role in the regulation of high temperature stress in Kareliniacaspia.At the same time,a large number of differentially expressed genes with certain high temperature resistance were obtained,which provided basic data for revealing the high temperature molecular mechanism of Karelinia caspia.The overexpression vector of growth hormone synthesis related gene Kcb HLH71 was constructed to lay a foundation for further verification of the function of this gene in response to high temperature stress. |
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