Cloning Of Cucumber LCD And DES Gene And Their Response To Abiotic Stress

Hydrogen sulfide?H₂S?,an important gas signal molecule,which could be generated endogenously in both animals and plants.H₂ S is involved in a range of physiological processes in plant,including growth,development,as well as biotic and abiotic stresses resistance.Enzymatic activity assays showed that L-cysteine desulfhydrase?L-CD?was primarily responsible for the generation of endogenous H₂ S in plant.As an important vegetable crop,cucumber is rich in vitamins.The determination of cucumber L-CDs is helpful for understanding the stress resistance mechanism of cucumber,which would provide some guidance for cucumber resistance resources breeding.This paper studies the response of endogenous H₂ S signal to high temperature?37??,low temperature?4??,osmotic stress?PEG8000?and low light?70% shading?environment,and identified two activated L-CDs from cucumber 9930 varieties.The results are as follows:?1?H₂S content kept higher than control when cucumber was treated by 4? or PEG 8000 from 2 to 8 hours,or treated by 37? or low light from 4 to 8 hours.H₂ S production rate rised after 37? or low light treated for 4 hours,or PEG 8000 treated for 2 hours.In the case of 4? treatment,the H₂ S production rate rised when plant were treated about four hours.?2?The Csa LCD gene responded to high temperature,osmotic stress,and low light conditions.The m RNA relative level of Csa LCD was lifted under 37? from 2 to 8 hours,or under PEG 8000 treatment for 4,6,or 8 hours.It was also rised under low light condition for 2 hours,and then back to the level of control.The Csa DES gene responded to high and low temperature stress.After 2 hours of 4? treatment and 4 hours of 37? treatment,there was a significant enhancing in Csa DES 2 expression.?3?Three L-CDs genes,Csa LCD,Csa DES1 and Csa DES 2 were acquired by sequence alignment.The CDS sequences of Csa LCD?Csa 2P034800.1?and Csa DES 2?Csa1P574810.1?were amplified successfully by PCR;The recombinant proteins His-Csa LCD and His-Csa DES 2 exhibited evident L-CD activity.?4?The CDS of Csa LCD or Csa DES 2 with 35 S promoter were transformated into lcd/des1 mutant of Arabidopsis,and both their heterologous expression restored the H₂ S production rate and restored the the mutant stomata's that is insensitive to PEG8000 treatment.This study showed that Csa LCD and Csa DES2 are the main enzymes for endogenous H₂ S production in cucumber,and there are some differences in response to stress.