CDNA Isolation And Function Study Of Aquaporin CpTIP From Chimonanthus Praecox(L.) Link

Frost, drought, salinization of the soil or acid, high temperature, high light illumination or inadequate environmental stress can affect the normal growth of plants, directly restrict the output of crops, and hamper the development of agriculture. In all of these environmental stress, the stress caused by lack of moisture most widely exists. Water shortage is not only weather-related drought, but high concentration of salinity can result in plant tissue infiltration of balance, so that cell water loss, thereby indirectly cause plant arid environment within the organization. Aquaporin (AQP) is commonly found in the biomembrane of animals, plants and micro- organisms. It is a kind of specific and high efficient water-transporting channel through cell membrane system. The rate of transmembrane water flux may be controlled by changing the abundance or the activity of the aquaporins.Chimonanthus praecox (L.) Link has typical treats of resisting drought and freeze. We used Clontech the Creator SMART cDNA Library Construction Kit to successfully construct Chimonanthus praecox (L.) Link full-length cDNA library. Original library titer was 1.16×105 cfμ/mL and recombinant rate was 96.43%. We cloned the open reading frame (Open Reading Frame, ORF) sequence, including 753 bp and encoding 251 amino acids. its pI is 6.14. Through BLAST and evolutionary homology analysis of the amino acid sequences for bioinformatics analysis, we find a high content of hydrophobic amino acids, and six transmembrane regions.and also find the gene contained the typical signal sequence SGXHXNPAVT of MIP family.Its B and E Ring has aminoacyl-proline-alanine motif (NPA) which are landmark structure of AQP family members. By phylogenetic tree analysis, it is close toγ-TIPs of the peach and pear. So this gene should belong to TIP sub family of AQP family andγ-TIP subclasses, and named its CpTIP.In order to study their biological function, we constructed plant expression vector pTEV7::CpTIP. With Agrobacterium transformation of Arabidopsis thaliana anther infection and screened positive strains on Ms medium containing hygromycin from T0 generation seed, Finally got 41 transgenic plants. These plants were cultivated under salt (NaCl concentration of 50 mM, 200mM, 300mM), alkaline (pH 7.0, 8.0, 9.0), drought (PEG6000 concentration of 10%, 20%, 30%) and cold (4℃training 60hr) stress, then determinated the biomembrane conductivity of leaves and observed phenotypic changes. According to the same level of treatment to transgenic plants and wild-type Arabidopsis thaliana conductivity of the results of the comparison, we found the salt-tolerance ability and drought-tolerance ability of transgenic Arabidopsis were improved than wild type. In addition, it shows the possibility Characteristics of cold-tolerance.In order to know more functions of the CpTIP, we needs further studies in future.