Identification Of The Function Of Polygalacturonase Gene CaPG2 During Fruit Cracking In Capsicum Annuum L.

Capsicum(Capsicum annuum L.)is rich in nutrients.It is one of the indispensable seasonings in our daily life.It has a good health care effect on our human body and can help the human body to invigorate the stomach and eliminate food,promote blood circulation and reduce swelling.However,peppers are prone to cracking during fruit ripening or development,which has a serious impact on the yield and quality of peppers and will bring us huge economic losses.Fruit cracking is closely related to the senescence of fruit tissues and the changes in the structure of the cell wall of the peel.Active oxygen metabolism is an important cause of tissue senescence.Changes in the structure of the cell wall will affect the strength of the cells,leading to water imbalance and leading to fruit cracking.Glycoside hydrolase 28 family(GH28)members can hydrolyze pectin,change cell structure,reduce cell wall strength and disintegrate cell wall structure.The tomato fruit splitting gene SIPG and Arabidopsis fruit splitting gene ADPG1 were compared in pepper GH28 family to obtain the most homologous CaPG2 gene,and the expression of CaPG2 gene reached the highest level in the severe fruit cracking stage of pepper growth and development.It is speculated that CaPG2 gene is involved in pepper Regulation of fruit cracking.Therefore,we studied the enzyme activity and cell structure of the easy cracking pepper strain ‘SY06’ at different fruit cracking stages,and analyzed the function of the pepper GH28 family member CaPG2 gene on pepper fruit cracking.The following main results were obtained:1.The anatomical structure of the fruit of the easy-cracking pepper strain’SY06’ in the early,middle and severe stages of fruit cracking found that as the degree of fruit cracking in SY06 fruit increased,the stratum corneum gradually became thinner and the shape of the epidermal cells changed.Irregularities of the sub-epidermal layer,increased spacing between cells in the sub-epidermal layer,and inconsistent arrangement of cells and cells;researches on the antioxidase and cell wall hydrolase enzymes in the early,middle and severe stages of fruit cracking in the fragile pepper strain’SY06’;As the severity increases,the activities of SOD,CAT,POD,PG and CX gradually increase,APX activity gradually decreases,and β-Gal activity first increases and then decreases.2.Analyze the expression pattern of the CaPG2 gene using the easy-cracking pepper strain’SY06’: the CDS of the CaPG2 gene is 966 bp in length,encoding 321 amino acids,the isoelectric point is 5.69,and the molecular weight is 35.23KDa;under normal conditions,CaPG2 is in the cracking of pepper fruit.The expression level in severe and mid-term is higher than that in the previous period.The transient expression in tobacco proved that the CaPG2 protein was localized in the cell membrane and nucleus.It shows that the CaPG2 gene is involved in the regulation of pepper fruit cracking.3.After using VIGS to silence CaPG2 gene in pepper fragile strain’SY06’,it was found that the CaPG2 expression level of pepper after silence was significantly lower than the control,and the silence efficiency reached 95%.The silent fruit had a lower fruit cracking rate than the control fruit.Higher hardness,denser stratum corneum and tighter cell arrangement;the activities of silent fruit SOD,POD,PG,CX and β-Gal are lower than the control,and the activities of APX and CAT are higher than the control.4.Using the Agrobacterium-mediated method,overexpression of CaPG2 in Micro-Tom found that the CaPG2 gene expression of transgenic fruits was significantly higher than that of WT;after soaking the transgenic fruits and WT with 2.5g/LCa Cl2,it was found that the CaPG2 gene was transfected.The fruit has a thinner stratum corneum,a larger cell spacing and a looser cell arrangement;the transgenic fruit has higher SOD,POD,PG,CX and β-Gal activities than WT,and lower APX and CAT activities.