Cloning And Fuction Identification Of CgHSP70Gene From Chrysanthemum Grandiflorum

Heat shock proteins (HSPs) comprise a protein family whose expression can be induced when a cell is responding to high temperature, salinity, drought, starvation and heavy metal etc enviromental stress. Functioning as molecular chaperones, HSPs can prevent the accumulation of protein precursors, accelerate the transport of proteins, and absorb complexes of unfolded proteins to maintain their transport abilities. HSPs can also sustain the normal folded state of proteins, degrade misfolded proteins, stabilize polypeptide strands, and prevent protein inactivity. It plays an important role for plants to overcome disadvantage condition.Ground-cover chrysanthemum’Zhongshanzigui’is an excellent species because of its high temperature tolerance. In this study, a full length cDNA of heat shock protein70gene named CgHSP70was isolated from chrysanthemum grandiflorum by means of RT-PCR and RACE, primers used in this experiment were designed based on the conservation of sequence from EST of our laboratory. The full length cDNA of CgHSP70was2205bp, and the ORF was1947bp which was predicted to encode a polypeptide of647amino acid residues. CgHSP70showed a highly identity with SmHSP70, AaHSP70and NtHSP70, come from Saussurea medusa, Ageratina adenophora and Nicotiana tabacum, and the identities were98.92%,98.77%and93.3%. Phylogenetic tree analysis showed that CgHSP70belonged to heat shock protein, not heat shock cognate protein, and it has a close relationship with AaHSP70.Transgenic CgHSP70Arabidopsis thaliana showed enhanced tolerance to high temperature, drought and high salinity, and the survival rate of transgenic plants nearly to90%, while the survival rate of untransgenic plants were all zero. The analysis of Quantitative real-time PCR showed that expression of CgHSP70in transgenic plants under stress condition is higher than normal condition.The result of transgenic chrysanthemum showed that CgHSP70has successfully transformed into chrysanthemum cutting cultivar’Jinba’, The survival of transgenic plants, transformed empty vector plants and WT plants under heat shock stress is100%,0and0respectively, under drought stress is100%,40%and45%respectively, and under salinity stress is100%,80%and75%respectively. The transgenic showed enhanced tolerance to hot, drought and salt stress. The expression of CgHSP70in transgenics at most is3.14, folds,3.10folds and4.88folds than contro under heat shock, drought and salt stress, respectively. In addition, the semi-lethal high temperature (LT50) of three transgenic lines, vector and WT plants is50.20℃、50.31℃、50.15℃、47.99℃and47.71℃respectively, the LT50of transgenic seedlings is significantly higher than control. Physiological index such as POD, proline and malondialdehyde are all detected, the cotent of proline and POD activity of transgenic plants are all higher than control under high temperature, while the content of malondialdehyde is lower. The result showed that the transgenic plants may be acquired the oxidation and permeation resistance. The POD activity, content of proline and malondialdehyde showed same tendency under drought and salinity stress conditons.