Characteration Of SlCBL2 From Solanum Lycopersicum

Calcium ions(Ca2+)act as one of the leading secondary messenger in the plant signal transduction pathway and play a fundamental role in responses to multiple extracellular stimuli,such as biotic and abiotic stresses,and regulating various cellular and developmental processes.Cellular calcium signals including distinct spatiotemporal changes in Ca2+concentration are detected and transmitted by sensor molecules such as calcium-binding proteins.It is well document that CBLs-CIPKs signaling systems are required for tolerating stresses in Arabidopsis,however,it remains elusive in tomatoes.Therefore,it is necessary to study CBLs-CIPKs associating with stresses and development in tomatoes.On the basis of transgenic Nb over-expressing and RNAi SlCBL2,its function was characterized.The results were as follows:Transgenic plants in seed,seedling and adult stages were treated by salt,low potassium,high pH and oxidative stresses.Salt,low potassium and oxidative stresses suppressed germination of all the genotype seeds,however,rates of germination among 3 genotype seeds were not statistically difference.High pH stress little affected germination,but growth of seedlings was repressed,compared with that of the over-expressing.Growth of all the genotype seedlings was similarly inhibited by salt and low potassium stresses,however,lines over-expressing increased tolerance to high pH and oxidative stresses.Salt,low potassium and oxidative stresses suppressed the growth of adult plants,but not statistically difference among three genotypes.On the contrary,over-expressing adult plants statistically enhanced tolerance to the high pH stress.Unlike wild type plants,optimal/maximal photochemical efficiency of PS II in the dark(Fv/Fm)in over expressing plants decrease little under the high pH stress.The improved BiFC vectors,p1301-nEYFP-c/cEYFP-c,has been successfully used,however,higher expression of target gene,which is driven by double 35s promoter,leads to increase false positive rate.To avoid the drawback,we re-improved BiFC vector on the basis of 1301 and acquired 1301-nEYFP/cEYFP,which have capability inserting promoter and target gene.Interactions of SlCBL2 driven by native promoter and 35s with five SICIPKs(SlCIPK3?SlCIPK7?SlCIPK8?SlCIPK9?SlCIPK11?SlCIPK23?SlCIPK24),which were driven by 35s promoter,were screened,suggesting that pCAMBIA1301-pSlCBL2-nEYFP-SlCBL2 only interacted with SlCIPK8.However,SlCBL2 driven by 35s can interact with four SICIPKs including SlCIPK3,SlCIPK8,SlCIPK11 and SlCIPK23.Therefore,we thought re-improved vector succeed in decrease false positive rate and SlCBL2 interacted with SlCIPK8,but not SlCIPK11.To furtherly investigate the available cues of SlCBL2 increasing tolerance to high pH stress,expressing profile of down stream genes is detected by real-time PCR.NbNHX2 and NbAHA2 were constitutively activated by SlCBL2;expression of NbDREB2 and NbVHAl is not associated with SlCBL2 and high pH stress;...