| Helianthus tuberosus L., a perennial herb,is rich in chrysanthemum sugar.It has many Benefits such as frequent with high precipitation,The acid rain is high in red soil region of southern China.Red soil generally exhibits strong acid reaction, And is rich for aluminum. Aluminum toxicity should be considered when elianthus Tuberosus extends its panatation to Southern of China. To study on growth of Helianthus Tuberosus under aluminun stress mechanism will provide scientific and practiced basis to improve yield and find the methods to relief aluminum stress.The ecophysiological response of Helianthus tuberosus L.To acid-aluminum stress and effect of exogenous Salucylic acid(SA) was shudied by plant physiolgical methods. Cysteine proteinase inhibitor gene was obtained from Helianthus tuberosus L. Leaves using RT-PCR. Bioinformatic was used to analysis the gene structure. In order to explore the relationship between cysteine proteinase inhibitor gene and stree, the gene expression under aluminum will be studied. The main results as follows:1. A solution culture experiment was conducted to study the effects on root growth, leaf photosynthetic characters, antioxidant enzyme activities, soluble sugar, and MDA content of two Helianthus tuberosus, Nanjing (aluminum tolerant cultivar) and Ziyang (aluminum sensitivity cultivar), under acid-aluminum stress. The result showed that the pH of solution and concentration of aluminum had distinct effects on physiology of Helianthus tuberosus. The normal growth of Helianthus tuberosus was affected by different pH ever under the minimal aluminum concentration. Compared with the control group, root length of Helianthus tuberosus was significantly inhibited, but there was little difference between the two genotypes. Root activity increased, while chlorophyll content, maximum energy conversion efficiency (Fv/Fm), and electron transport rate were significantly decreased in both genotypes of Helianthus tuberosus. SOD, POD, and CAT activity showed opposite trends. The inhibition on Helianthus tuberosus (Ziyang) was significantly less than that of Helianthus tuberosus (Nanjing), indicating the differences between the two genotypes in aluminum tolerance. Initial flurescence (Fo), soluble sugar, and MDA content were increased, while qP and qN showed opposite trends. With the increasing of aluminum concentration, the differences of physiological indicators between aluminum treated group and control group of Helianthus tuberosus increased. Under the same concentration of aluminum, the root activity increased, chlorophyll a decreased, Fo increased, soluble sugar content decreased, SOD activity and MDA content increased when it was under pH4 compared to pH5. Under different aluminum concentration, the physiological indicators of Helianthus tuberosus were significantly different. Roots activity and antioxidant enzyme activity were raised with increasing aluminum concentrat-ion. It can be deduced that the antioxidant enzyme system and root absorption mechanism played an important role in relieving acid and aluminum stress. Through the grey correlation analysis, the closest indicators with the acid and aluminum stress of Helianthus tuberosus was chlorophyll qN, followed by ETR, MDA, and Fv/Fm.2. A solution culture experiment was conducted to study the effects of exogenous salicylic acid(SA) of 50μmol/L on growth characteristics, antioxidant content and antioxidant enzyme activity of two Helianthus tuberosus cultivars, Nanjing (Al resistance genotype) and Ziyang (Al sensitive genotype) under Aluminum stress. The result showed that:the Helianthus tuberosus seeding under 500μmol/L Al stress for 21days,the plant height, root length and fresh shoot weight, The chlorophyll content,the maximum energy conversion efficiency(Fv/Fm). maximum fluorescence yield (Fm) and electron transport rate (ETR) were significantly lower of two Helianthus tuberosus SOD, POD and activity changed in the opposite trend. Indicating the differernce of the two Helianthus tuberosus. Initial fluorescence (Fo),the contents of soluble protein,O2- production rate, proline (Pro) and MDA increased. Spraying exogenous salicylic acid(SA) of 50μmol/L improved the root length,The chlorophyll content, antioxidant enzyme;reduced the O2- production rate. the contents of soluble protein,proline (Pro) and MDA. Basedon on the analysis of membership function,Aluminum by can alleviate the damage of Helianthus tuberosus， Spraying 50μmol/L SA every day enhanced the ability of Helianthus tuberosus to resistance. It can be concluded that Spraying SA influence chlb,Fm and ETR of Helianthus tuberosus by the grey correlation degree analysis.3.Cysteine proteinase inhibitor(CPI) is a gene family that widely exist in plants, and it mainly involved in plant plastid development, growth and resistance, the open reading frame of CPI was cloned from Helianthus tuberosus L.eafs with RT-PCR. The expression of CPI under stress wsa analysised and using bioinformatics to analysis the gene. The result showed that the sequence contains a characteristic domain of CPI. HtCP1 was 333bp, contains 4 Glycosylation sites and 2 Protein kinase C phosphorylation site. The protein structure prediction showed that HtCY contanis transm embrane and signal peptide. Phylogenetic analysis indicated that HtCY had high identity (94%) with Curcuma longa L. The amino acid alignment result deduced HtCY contains a high conserved QKVKQ domain and a conserved LAKFAVDHHN domain. The POST result showed, HtCY may locate in the endoplasmic reticulum. The tissue specificity tests showed that HtCPI expressed in roots, stems, leafs, and it has high expression in stems. The Real-time PCR results indicated that HtCPI expression was enhanced under AlCl3,NaCl,PEG,SA stressand has the highest expression at 6h,12h,24h. It shows that the gene is involved in the defense mechanisms of Helianthus tuberosus to biotic stress. |
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