Functional Analysis Of SlRCHY3 Gene In Tomato (Solanum Lycopersicum)

Tomato is a kind of vegetable with high nutrition,small genome,short growth period,mature transformation system and complete genome sequencing.The RING domain containing zinc finger protein is also named RING type zinc finger protein.Most of the E3 ubiquitin ligases contain RING domain,and most of the RING type zinc finger protein can produce to participate with other proteins involved in ubiquitin protein degradation,gene expression,signal transduction and transcription regulation of a series of biological reaction.In this study,Sl RCHY3 gene was cloned and its expression pattern was detected.We explored its biological function by constructing over-expression vector and transforming it into explants of Wild type tomato.The main results are as follow:(1)Sl RCHY3 gene encodes 280 amino acids.The protein contains RING domain,and it is concluded that the protein encoded by Sl RCHY3 may be RING type zinc finger protein.(2)The expression pattern in Wild type tomato tissues showed that the highest expression level was in old leaves,roots while low expression in fruits.The upward trend,expression appeared in young leaves,mature leaves and old leaves which indicates that Sl RCHY3 gene may involved in the development of leaves.(3)We detected the expression level of Sl RCHY3 under the treatment of abiological stress and methyl jasmonate.In Results showed that the expression of Sl RCHY3 gene was induced under the treatment of low temperature,high salt and drought significantly,which suggest that Sl RCHY3 gene may involved in abiotic stress control process.There was no significant change in the expression level under methyl jasmonate treatmentwhich indicating that the expression of Sl RCHY3 gene might not be affected by methyl jasmonate.(4),Sl RCHY3 gene expression vector was constructed by the application of molecular biology techniques and transformed into Agrobacterium.Tomato cotyledons were infected with Agrobacterium by agrobacterium-mediated gene transformation Sl RCHY3 overexpression transgenic lines were screened.And according to the observation and statistics,fruit did not change significantly.(5)We found that in transgenic tomato plants,leaf senescence was slower than the Wild type.And increased chlorophyll content.Through the detection of senescence related genes,photosynthesis related genes Sl RBCS and Sl Cab7,the expression of chlorophyll synthesis related genes DCL,Golden2like1 gene expression level in no treatment of transgenic lines than the Wild type is still higher than the wild-type in days.Sl RCHY3 gene on chlorophyll synthesis showed positive regulation,no expression of Sl Cab7 and Sl RBCS genes emerged significantly higher than that of the Wild type,after processing is still higher than the Wild type,Wild type photosynthesis weakened,accelerate leaf senescence over expression of Sl RCHY3 gene,can delay leaf senescence.In MS medium,the length of lines of over expression significantly lower than the Wild type,we speculate that overexpression of the gene can inhibit the development of root,wild type and overexpression lines hypocotyl length is basically the same.Indicating that the Sl RCHY3 gene does not affect hypocotyl development.(6)In the T1 positive strain,the dehydration and high salt treatment experiments were carried out,and the leaf water content was detected.The results showed that the Wild type AC was higher than that of the transgenic lines.The water loss rate of AC was lower than that of the transgenic lines.The results showed that the expression of abiotic stress related genes in transgenic lines was lower than that in Wild type.In the high salt stress,we found that the transgenic lines were wilting and dead,and further detected the abiotic stress related genes,and found that the expression level of AC was higher than that of the transgenic lines.These results suggest that overexpression of Sl RCHY3 gene can reduce the drought and salt resistance of plants.