Cloning Of The PlSAURs Genes From Paeonia Lactiflora Pall. And Transformation Of Arabidopsis Thaliana

In this study,four full length cDNA of PlSAURs genes were cloned from different developmental stages(PIB,before the imbibition;PIA,after the imbibition;PDA,after germination of down-hypocotyl;PUA,after germination of up-epicotyl)of Paeonia lactiflora hybrid seeds(?“Fen Yunu” × ?“Fen Yulou”).By the agrobacterium-mediated method,the constructed plant expression vector was transferred into Arabidopsis thaliana.PlSAURs genes were transformed into Arabidopsis thaliana by agrobacterium-mediated transformation system,which lay a strong basic for further understanding the effect mechanism of PlSAURs genes in dormancy and germination.The main results of this study are showed below:1.Total RNA was extracted from four different key developmental stages of Paeonia lactiflora hybrid seeds respectively.The levels of PlSAURs genes expressed by Real-Time fluorescence quantitative RT-PCR.The results indicated that PlSAUR1 was abundant in PDA stage,PlSAUR2 was abundant in PUA stage,PlSAUR3 was abundant in PUA stage,PlSAUR4 was abundant in PIA stage.It is also laied a theoretical foundation that select the most optimal period of P.lactiflora seeds as PlSAURs genes cloning plant materials.2.PlSAURs genes were cloned by PCR from P.lactiflora seeds successfully.The results indicated that the PlSAUR1 full length open reading fragment was 495 bp in length,encoding 164 amino acid residues;the PlSAUR2 full length open reading fragment was 381 bp in length,encoding 126 amino acid residues;the PlSAUR3 full length open reading fragment was 300 bp in length,encoding 99 amino acid residues;the PlSAUR4 full length open reading fragment was 558 bp in length,encoding 185 amino acid residues.The amino acid sequences of the PlSAURs were compared with the amino acid sequences in the NCBI database.The high similarity and coverage homologous sequences were selected for homology and phylogenetic analysis.And the PlSAURs protein structure and other bioinformatics were analyzed by online software.3.Inserting the coding region of PlSAURs genes to plant expression vectors of pBI121,and then transgene to Arabidopsis thaliana by Agrobacterium tumefaciens.Transgenic plants were tested at DNA and RNA levels.The results indicated that PlSAURs genes have been transformed into Arabidopsis thaliana successfully.