Cloning Of The PLPP2Cs Genes In Paeonia Lactiflora Pall.and Transformation In Arabidopsis Thaliana

Peony(Paeonia lactiflora Pall.)is a perennial herb flower,which has a long history of cultivation and is one of the traditional famous flowers in china.The flowers are colorful and varied,and are now widely used in the field of landscaping,which can be used as special gardens,flower beds and flowers.The combination of tree peony and peony can extend the flowering period at the visual level and improve the viewing effect.However,the seed has the dormancy of the upper and lower embryo shaft,which hinders the breeding and production work of the drug,and causes the peony to not meet the demand of the ornamental flower market completely.In this study,the hybrid seeds of the peony cultivars “Fen yu nu” and “Fen yu lou” were used as materials.Differentially expressed genes Pl PP2 Cs were screened based on transcriptome data during the germination of Paeonia lactiflora seeds obtained earlier by the research team.Cloning obtained the full length of the gene and carried it bioinformatics analysis,using real-time fluorescence quantitative PCR technology to analyze the expression of The Pl PP2 Cs gene in different periods of germination of the seeds of Paeonia lactiflora,and constructed the p BI-Pl PP2 Cs plant expression vector,after the transformation of agricultural bacteria,genetic transformation of wild type Arabidopsis thaliana,The resistant Arabidopsis thaliana was obtained through antibiotic screening and PCR molecular identification was obtained to lay the foundation for verifying the function of the Pl PP2 Cs gene.The results of this study are as follows:1.According to the transcriptome data obtained by the research team,the differential expression gene Pl PP2 Cs were screened,and the total length of the gene was obtained.Pl PP2C1 gene ORF is 1236 bp in length and encodes 411 amino acids.Pl PP2C2 gene ORF is 1341 bp in length and encodes 447 amino acids.The Evolution Tree of the Pl PP2 Cs Protein System shows the recent kinship between Pl PP2C1 and the PP2 C of Eucalyptus Grandis.Pl PP2C2 is close to the PP2 C of Vitis vinifera and Nyssa sinensis.The proteins encoded by Pl PP2C1 and Pl PP2C2 belong to the hydrophilic,non-transmembrane unstable proteins,and both contain PP2 C protein conservative domains.Subcellular location prediction results show that both The proteins encoded by Pl PP2C1 and Pl PP2C2 are the most likely to be positioned in the nucleus.2.After extracting the total RNA from different periods of germination of the seed,the expression of the Pl PP2 Cs gene in different periods was analyzed by using real-time fluorescence quantitative PCR technology.The results showed that the expression of The Pl PP2C1 gene increased after inflation and whitening,and decreased gradually from the beginning of rooting,indicating that the Pl PP2C1 gene helped to break the perication of the embryo shaft under the drug.The expression of the Pl PP2C2 gene in the stage of erythema and long buds in the roots increased significantly,and it is assumed that it contributes to the long buds of the sage seeds and has a catalytic effect on breaking the upper embryo shaft sleep.3.Using homologous recombination to construct p BI-Pl PP2 Cs plant expression vector,using the frozen melt method to successfully transform the expression vector into agricultural bacteria,the wild type of amoeba genetic transformation.The resistant Arabidopsis thaliana was obtained by resistance screening.