Functional Analysis Of AM-dependent AsPT1and AsPT4Gene Upstream Acting Elements

Mycorrhizal symbiosis is a widely present in terrestrial plants, and works with more than80%of terrestrial plants form a symbiotic relationship, the absorption of N, P, Fe, Cu and other elements of the plant have promoted, especially in the P element absorption is more prominent, fossil analysis shows that this symbiotic relationship exists once in460million years ago. When the plants and arbuscular mycorrhiza fungi (AMF) formation symbiosis, the plants have acquired a new P absorption pathway that arbuscular mycorrhizal fungi pathway or the indirect pathway.through this pathway, plants not only improve the P absorption efficiency, but also expanded plants absorption area of P in soil. And studies have proven that indirect absorption pathway plays a key role in P absorption of AM plants (Smith et al.,2003,2004).Although the role of this symbiotic relationship has been widely accepted, however, the molecular mechanism of this symbiotic relationship is not clear. We are focus on the regulation of phosphate transporter genes (PTs) expression which associated with the AM. Our laboratory have been isolated six P transporter genes, according the expression pattern of the six PTs gene, we choose two kinds of mycorrhiza-specific P transporter genes that AsPTl and AsPT4, based on phylogenetic tree analysis showed that both belong to Ph1phosphate transporter family; RT-PCR and RT-qPCR analysis confirmed AsPTl and AsPT4is mycorrhizal specific genes again, is expression only in the AM symbiosis. Using bioinformatics methods detect the putative motif inside promoter region of AsPTl and AsPT4gene, as well as FootPrinter2.0prediction new motif both of them. For vector construction using PCR and over-lap PCR method, using Agrobacterium K599-mediated methods for plant transformation, transformed plants were co-cultured with Glomus intraradices four weeks. TB staining, GUS staining and detection of GUS activity of the transformation roots in the final. The result is as follows:1, As mycorrhiza-specific phosphate transporter gene cis-acting elements MYCS also has an important role in regulating of mycorrhiza special phosphate transporter gene AsPT1and AsPTl of Astragalus sinicus2, The regulation of AsPT1gene expression is more complex, the TGTT-motif, AAAA-motif, TAAT-motif, MYCS and other components within of AsPT1promoter can play as a cis-regulatory modules (CRMs), there is also presence both positive and negative regulation. TGTT-motif elements play a significant role in regulating of AsPTl gene.3, The regulation of AsPT4gene expression is relatively simple, the TAAT-motif and TAAC-motif can increase the expression distinctly, possibly as an enhancer element.In summary, TGTT-motif, AAAA-motif, TAAT-motif and TAAC-motif of the mycorrhizal specific phosphate transporter gene AsPTl and AsPT4of Astragalus sinicus has obvious role in expression regulation. The orthologous phosphate transporter gene promoter sequence analysis showed that these element found in this article are widely present in mycorrhizal phosphate transporter gene promoters, they may be have a similar function with the MYCS, can regulate most of AM-dependent PTs...