Cloning and functional analysis of the promoter of allergen gene Ara h 1 from peanut

Peanut seeds are ideal bioreactors for the production of foreign recombinant proteins or secondary metabolites.Seed-specific promoters(SSPs)can direct the expression of genes specifically in seeds to avoid undesirable effects associated with constitutive expression.However,few SSPs have been identified in peanut.Previous studies have shown that some allergen-encoding genes encode seed storage proteins or exhibit seed-specific/preferential expression.In this study,we characterized allergen-encoding genes from across the genomes of Arachis species to explore seed-specific genes.We found that at least 9 out of 16 identified peanut allergen-encoding genes were expressed specifically in the seeds or were preferentially expressed.A 1493-bp promoter fragment of allergen gene Ara h 1(we named it AHSSP6)was isolated from cultivated peanut genome.cis-element analysis showed that three RY repeat elements which usually exsisted in seed or embryo specific promoter sequence were also present in AHSSP6 sequence.Histochemical analysis showed AHSSP6 could drive the expression of aβ-glucuronidase(GUS)reporter gene specifically in the seeds or cotyledon tissue of transgenic Arabidopsis,while not in other tissues.These findings indicated that these promoters of allergen genes were candidate SSPs,and AHSSP6 was a novel SSP which could be potentially utilized in peanut improvement.

Cloning and Functional Analysis of Xylem Specific Promoter Deletion Fragment

Xylem-specific promoter could regulate efficient expression of foreign genes in xylem. Deletion analysis method was applied to obtain 5＇ deletion promoter fragments MU2 andMU4 with different distances from transcription start site of xylem-specific promoter MDCcsAP through PCR procedure. Then CaMV35S promoter in plant expression vector pBI121 was replaced by amplified fragments. The recombinant plasmids fused with corresponding gene segments and GUS reporter gene were obtained and transformed into Agrobacterium. The results of the tobacco transient transformation test showed that MU2 andMU4 had the promoter function, and both of their activity were significantly higher than that of CaMV35S promoter, and they had the characteristics of xylem tissue specificity as well. Research on xy- lem-specific promoter structure and function could lay a foundation for the determination of necessary components to the xylem tissue specificity and cis-acting ele- ments with induction activity, so as to enable the regulation of efficient expression of exogenous genes in specific time and specific tissues in plants through these cis-acting elements.