| Rosa rugosa is a deciduous shrub which belongs to Rosa genus of Rosaceae family.The shrubs having splendid color and delightful fragrance are cultivated in many varieties. It is not only a excellent plant used in landscape greening and water and soil conservation project, but also a valuable material used in Perfume and food industry.Wild Rosa rugosa originates from china and is one of the state second class protected species. It mainly distributes in Jilin Tumen River Estuary, the south coast of Liaoning and the north coastal area of Shandong. Because of the survival environment,wild Rosa rugosa has strong salt resistance, and become valuable material of breeding new variety. It is reported that the vacuolar Na+/H+antiporter and vacuolar H+-ATP protease play important roles in plant salt tolerance. Vacuolar Na+/H+antiporter (RrNHXl) and vacuolar H+-ATPase subunit c (RrVHA-c) gene of wild Rosa rugosa from weihai of Shandong province were cloned and expression of gene was analyzed in this study, it has important academic significance. The main results are as follows:Total RNA from young leaves was extracted using the CTAB method, then the full sequence of gene RrNHXl and RrVHA-c gene which related to the salt resistance were cloned by RACE method. The result for the test shows that RrNHX1CDNA contains2358nucleotides with an open reading frame of1629nucleotides and was submitted to GenBank with accession number KC188664.The gene encodes a543amino acid protein with deduced mass of60.10KD.The protein was composed of a hydrophobic N-terminus,10transmembrane helix regions and2non-transmembrane helix regions, and a hydrophilic C-terminus. The third transmembrane helix region contains the binding site of amiloride.The RrNHX1belongs to the type of NHX-I family located in vacuole membrance,which far from the type of NHX-â…¡ family located in membrane vesicles and further from the type of SOS1family located in plasma membrane. The result also shows that RrVHA-c gene contains781nucleotides with an open reading frame of498nucleotides and was submitted to GenBank with accession number KF677023. RrVHA-c gene encodes a543amino acid protein with deduced mass of16.7KD.The protein was composed of5hydrophobic regions which contain4transmembrane helix regions. RrVHA-c belongs to the V-ATPase subunit c family, a membrane protein is conserved in evolution.Using the real-time PCR technology, the expression of RrNHXl and RrVHA-c in the young leaves were analyzed after the treatment of salt.The study shows that RrNHX1constitutively expressed in leaves,and its expression is induced by salt stress.Its expression increased first and then decreased with the increase of salt concentration, and has a time regulating effect. This demonstrates that RrNHXl may play a peculiar role in stress resistance of Rosa rugosa. The expression of RrVHA-c has characteriatic of housekeeping gene.The expression of RrVHA-c is similar to RrNHXl and increase in coordination with RrNHXl under certain concentration of salt,which is consistent with Na+/H+antiporter and vacuolar H+-ATP protease having synergistic effects against salt stress and give further evidence of theiy are closely related to salt tolerance of wild Rosa rugosa. |
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