Identification Of Wheat Materials Transformed With TaPHR1 Without Selectable Marker And Molecular Identification Of N-efficient And P-efficient Germplasm

Phosphate is one of inorganic nutrients required by plants. It is a vital component in many important compounds of plants, and also participating in a large number of physiological processes. In the soil, there are mainly indissoluble bivalent or trivalent phosphates, which cannot be utilized to leads to yield and quality reduction in wheat. The status quo can be solved by Phosphate fertilization, which not only cannot solve the problem of Phosphorus deficiency fundamentally, will create more problems like environmental pollution, resource waste and cost increases. And the fundamental letout to solve the problem of P-deficiency is tapping the potential of phosphorus uptake and utilization of wheat and cultivating new p efficient wheat materials from genetics and breeding.The first part of the materials were obtained by biolistic particle co-transformation,which contains TaPHR1 P-efficient gene and bar selection gene, and 3 transgenic wheat line only with the target gene were identified finally. The rest 466 materials from our lab were analyzed using molecular marker linked to N-efficient gene and SSR marker linked to N, Pefficient QTL. The main results are as following:(1) 2 bar free wheat plants of 486 T1 transgenic wheat plants were obtained. 231 T2 transgenic wheat lines were identified and 3 lines only with target gene, which were from the same T1 plant P51, were obtained finally.PCR products sequencing proved that the TaPHR1 gene detected was exogenous. To resolve high false positive rate in transgenic bar wheat, PCR, smearing Basta on the leaves of transgenic wheat and lateral flow strip test were performed in this study, and the results shows that bar gene test could be accurately performed through PCR analysis in sterile condition, smearing Basta on the leaves of transgenic wheat with suitable concentration and lateral flow strip test.(2) The relactive expression of TaPHR1 gene in parent and transgenic wheat plants were analyzed by relative fluorescence quantitative PCR. The results indicate that TaPHR1 gene was highly expressed in transgenic wheat transformed with TaPHR1 gene, and the relative expression level was between 12 and 63.Dry matter accumulation and root shoot ratio of transgenic offspring materials of into TaPHR1 gene were higher than that of parents in hydroponics, and the differernce was particularly remarkable, which showed that TaPHR1 gene expression improvement resulted in stronger lower-phosphorous tolerance of transgenic offspring materials.(3) It showed that plant height variation and leaf color of transgenic offspring materials emerged while fertility was not remakable different with parents through investigation and analysis of transgenic offspring materials’ charactors.It was no difference of genome scanning results via SSR primers between parents and transgenic offspring materials from which we infered that cell variation caused by difference of exogenous gene insertion position results in plant height variation.(4) 466 varities or strains detection were performed through molecular makers of nitrogen and phosphorus high efficiency gene and SSR primers linked with nitrogen and phosphorus high efficiency gene QTL locus.It showed that authorized varieties possessed improved space and two materials possessed all efficient genes and QTL locus.