Temporal And Spatial Expression Of Rooting Gene PsARRO-1in Tree Peony

On the basis of cDNA full-length sequence PsARRO-1gene (GQ330644) which has beenextracted, the research studied the expression of ARRO-1using realtime PCR in different growthperiod and different part of ’Fengdanbai’ seedlings in field and sterile plantlets that were inducedby axillary bud. The quantitative analysis results of the gene expression will lay the foundationfor further study to investigate the gene function of ARRO-1. The results are as follows:1．Rference gene screeningFive housekeeping genes in the’Fengdanbai’, includingβ-Tubulin (EF608942), Actin,GAPDH,18SrRNA (U42792) and β-Actin were investigated using q-PCR. All data were collectedand analyzed by geNorm. β-Tubulin is chosen to be the proper housekeeping gene in thefollowing studies, for it shows specific and high efficiency of amplification, and the expression isthe most stable.2．The Real-time PCR system establishedThe PCR program was performed with three steps. A gradient of annealing temperature wasset as follows:52℃、54℃、56℃、58℃、60℃. The number of cycles was set as36,38,40,42,44. The best PCR reActin system was determined as follows according to the take-off time of themelting curve and the amplification effect: denaturing at95°C for2min, annealing at95°C for15s and58°C for20s, extension at72°C for20s, repeated over40cycles. The products were stableexpressed with single peak and normal trend curve.3. PsARRO-1gene expression patterns of explorationThe ARRO-1gene was expressed in seedlings, test-tube plantlets’s root, stem, leaf andflower. It is of higher level in seedlings than in test-tube plantlets. In the test-tube plantlet, thisgene is stable but the lowest expressed in stem, while it is only expressed in the first10-day in leafand it is of the highest level in root with tow significant peaks. In the seedlings, this gene is higherin root and stem with significant variance. The expression is moderate in leaf while it is lower inflower. Furthermore, the ARRO-1gene reached the peak at the10th day in leaf and root, while thepeak time is the35th day in stem. These results are corresponding with the prospective thatARRO-1gene modulates the adventitious root formation via regulating endogenous hormones.Further studies will be explored the physiological role making use of gene chip analysis.