Optimization Of The Genetic Transformation System Of Caladium Bicolor 'Hongtao K' And The Introduction Of AtPAP1 Gene And Maize Lc Gene

Caladium bicolor is a perennial herbaceous plant of the monocotyledonous family Araceae,and its leaves are often embedded with colored spots or colored veins.It is an important foliage plant suitable for greenhouse cultivation.Therefore,screening and breeding the new materials or new varieties of Caladium bicolor is an effective way to increase market competitiveness.Since anthocyanins play a major role in the formation of colorful leaf color in color leaf plants,the use of molecular breeding to modulate the synthesis of anthocyanins in color leaf plants is expected to rapidly acquire new materials or new varieties.Studies have shown that in the anthocyanin synthesis pathway,transcription factors directly regulate the transcription and expression of structural genes through the MYB-bHLH complex or the MYB-bHLH-WD40 complex?MBW complex?,thereby affecting the distribution and content of anthocyanins in plants.In this study,the genetic transformation system of Caladium bicolor‘Hongtao K'was optimized,and the plant expression vector pC1300-PAP1+Lc was introduced into Caladium bicolor‘Hongtao K'by Agrobacterium-mediated method.To explore the phenotypic changes of the transgene Caladium bicolor through the heterologous expression of the MYB type transcription factor AtPAP1 gene and the bHLH type transcription factor maize Lc gene.Provide technical support and theoretical reference for further landscaping and breeding.The main findings are as follows:?1?The genetic transformation system of'Hongtao K'was optimized.Using Caladium bicolor‘Hongtao K'as a material,the parameters related to genetic transformation were explored,and antibiotic concentration,antibacterial antibiotic concentration,preculture time,Agrobacterium infection time,co-cultivation time,concentration of total acetosyringone added to culture medium,negative Compressive strength,etc.,to obtain an optimized conversion system.The leaves of A were used as transforming receptors,pre-cultured for 3 days,and negative pressure was extracted 10times.100?mol/L acetosyringone?AS?was added when the OD₆₀₀00 of the inoculated bacillus was 0.3.The infection was 7 min,and co-cultured for 2 days in darkness at28°C.Adding AS to the inoculum and adding 100?mol/L AS in the co-culture can increase the genetic transformation efficiency of'Hongtao K'.At the screening stage,7.5mg/L hygromycin was added to the culture medium,and 500 mg/L carbenicillin was added as an antibacterial antibiotic.The transforming receptor was first placed into the callus induction medium,and the callus was formed and transferred to differentiation medium.After the resistant shoots grow out,they are transferred to the rooting medium for screening culture.The hygromycin-containing selection media must be replaced every 20 days.?2?Agrobacterium-mediated genetic transformation of AtPAP1 gene and maize Lc gene on Caladium bicolor‘Hongtao K'.In this experiment,Agrobacterium-mediated method was used to introduce the plant expression vector pC1300-PAP1+Lc into Caladium bicolor‘Hongtao K',and a new material was obtained.The phenotype of this material is obviously different from that of‘Hongtao K'.Its callus appeared red,the bud part was red,and the petiole and root of the seedling showed red.PCR detection identified that the gene of interest had been transferred to the transformed material.