Effects Of Piriformospora Indica On The Growth,Stress Resistance And Snakin Gene Expression Of Gerbera Jamesonii Bolus

Gerbera(Gerbera jamesonii Bolus),belonging to Chrysanthemum,is a perennial herb plant.It ranks the fifth in the global cut flower trade.Due to its self-incompatible characteristics,tissue culture propagation was widely used in Gerbera industry.However,long term subculture will weaken plant growth,which will influence the development of the gerbera industry.Moreover,the gerbera industry is threatening by root rot disease.Pirifomosporra indica has been proved to play important roles in promoting plant growth and development and stress resistance in many plants.Up to now,however,there is no report about the use of P.indica in gerbera.Therefore,in the present study,P.indica was used to identify its role in gerbera growth and development,stress resistance and root rot resistance.Additionally,we also investigated the effects of P.indica on the expression of gerbera Snaking genes(encoding antimicrobial peptides which were reported to play important role in plant-fungus interaction).The main results are listed as follows:1.Optimization of gerbera subculture and rooting mediumFor the production of high-quality gerbera seedlings,tissue-cultured Gerbera cv.Shuifen seedlings were used as materials for the optimization of the subculture and root medium.Results showed that MS+0.5 mg/L 6-BA+30 g/L sucrose+7.5 g/L agar(pH 5.8)was the best subculture medium and 1/2 MS+0.5 mg/L NAA+30 g/L sucrose+7.5 g/L agar(pH 5.8)was the best rooting medium for tissue-cultured Shuifen gerbera plantlets.2.Effects of Piriformospora indica on the growth and development of long-term subcultivated gerbera plantletsTissue-cultured Gerbera plantlets with different subcultivation times were used as materials and were respectively subjected to P.indica zymotic fluid,sterilized P.indica zymotic fluid and PDA liquid medium treatments.Growth indexes and chlorophyll content were measured at thirty and ninety days post treatment.Results showed that long term subcultivation weakened the growth of gerbera plantlets and lowered the seedling quality.Notably,P.indica zymotic fluid could promote rooting and growth of the tissue-cultured gerbera plantlets.3.Purification and identification of the pathogens causing Gerbera root rot disease in Sanming,FujianAccording to morphology and molecular identification analysis,Fusarium oxysporum and Phytophthora cryptogea isolates were purified from the diseased Gerbera plants showing root rot symptom in Sanming,Fujian.F.oxysporum and P.cryptogea were reinoculated to healthy gerbera plants.It was shown that P.cryptogea reinoculation could cause the root rot to gerbera.Although the F.oxysporum reinoculation could induce some diseased symptom but not the root rot symptom.And the co-infection of the two pathogens resulted in earlier symptom appearance and more serious symptom.Results generated in this part indicated that the root rot disease in Sanming,Fujian was mainly caused by P.cryptogea and F.oxysporum is helpful for its infection.4.Construction and optimization of the rapid detection method of the F.oxysporum in gerberaTwo pairs of LAMP-PCR specific primers and one pair of PCR-specific primers were designed according to the six sites of the mitochondrial partial sequence of Fusarium oxysporum for the rapid detection method of Fusarium oxysporum.The optimized detection system(total volume:25 μL)is:8 mmol·L-1Mg2+,0.8 mmol·L-1dNTPs,1.2 nmol·L-1 1-FIP/BIP primer,0.2 nmol·L-1 1-F3/1-B3 primer、0.4 mol·L-1 betaine,Bst DNA polymerase 8 U,10 × Reaction Buffer 2.5 μL and DNA 1.0μL;65℃,70min;85℃,10 min.5.Effects of Piriformospora indica on gerbera disease resistanceTo evaluate the effects of Piriformospora indica on gerbera disease resistance,the P.indica zymotic fluid,sterilized P.indica zymotic fluid and PDA liquid medium treated gerbera seedlings were co-inoculated with F.oxysporum and P.cryptogea.The highest POD,CAT and SOD activities and the lowest disease ratio were found in P.indica zymotic fluid treated gerbera seedlings,indicating that P.indica could increase the root rot resistance gerbera by regulating antioxidant enzyme activities.6.Effects of Piriformospora indica on the expression of gerbera Snakin genesBased on our gerbera transcriptome data,5 gerbera Snakin genes were obtained.By using quantitative real-time PCR,the effects of P.indica on the expression of gerbera Snakin genes were analyzed by comparing their expression levels in root rot pathogen inoculated gerbera seedlings that were P.indica colonized or not.Results showed that P.indica colonization influenced the expression of Snakin genes before and after root pathogen treatments,suggesting that the root rot resistance promoting effect of P.indica may be related to its influence on Snakin gene expression.