CONCLUSIONS: The small 295-bp ZmPht1; 5 promoter is sufficient to drive high-intensity expression of target genes, especially under phosphate deprivation conditions, and is therefore useful for crop improvement via transgenic techniques. Phosphate (Pi) deficiency has become a major challenge and limiting factor in world agricultural production. Manipulating the gene expression using appropriate promoters to improve the Pi absorption and utilization efficiency of crops could reduce the requirement for Pi fertilizers. In the study, a 295-bp strong promoter (M2P-7) of maize high-affinity phosphate transporter ZmPht1; 5 was isolated and functionally validated in transgenic Nicotiana benthamiana and maize by analyzing the ZmPht1; 5 promoter (M2P-1) and its 5\' truncated variants (M2P-2 ~ M2P-8) in different sizes under normal and Pi-deprivation conditions. The M2P-7 displayed the highest promoter activities among 5\' truncated fragments in all tested tissues of transgenic Nicotiana benthamiana at different development stages, which was 1.5 and 3 times higher than the well-used CaMV35S promoter under normal and Pi-deprivation conditions, respectively. In maize, the M2P-7 promoter activity was comparable to the maize ubiquitin1 promoter widely used in monocots under normal condition, which was about 1.3 times that of the ubiquitin1 promoter under Pi-deprivation environments. Moreover, the M2P-7 fragment is only 295 bp in length, thus reducing the construct size, and is therefore beneficial for genetic transformation. Thus, the small promoter M2P-7 of plant origin could be of great use for monocotyledonous and dicotyledonous crop improvement via transgenic techniques based on its promoter activities, expression patterns and small size.

A reliable pigeon pea transformation system can assist the rapid improvement of this important grain legume through transgenic development. Here we describe two methods of Agrobacterium tumefaciens-mediated pigeon pea transformation. In the tissue culture based embryonic explant transformation method, microshoot grafting was included to obtain rapid root induction, while the other method was culture independent and designated as plumular meristem transformation. Both methods drastically enhanced the transformation frequency and have the potential to provide reasonable solutions for maximum transgenic recovery in biotechnological breeding programs.

Transgenic technology has become an important technique for crop genetic improvement. The application of well-characterized promoters is essential for developing a vector system for efficient genetic transformation. Therefore, isolation and functional validation of more alternative constitutive promoters to the CaMV35S promoter is highly desirable.
In this study, a 2093-bp sequence upstream of the translation initiation codon ATG of AtSCPL30 was isolated as the full-length promoter (PD1). To characterize the AtSCPL30 promoter (PD1) and eight 5\' deleted fragments (PD2-PD9) of different lengths were fused with GUS to produce the promoter::GUS plasmids and were translocated into Nicotiana benthamiana. PD1-PD9 could confer strong and constitutive expression of transgenes in almost all tissues and development stages in Nicotiana benthamiana transgenic plants. Additionally, PD2-PD7 drove transgene expression consistently over twofold higher than the well-used CaMV35S promoter under normal and stress conditions. Among them, PD7 was only 456 bp in length, and its transcriptional activity was comparable to that of PD2-PD6. Moreover, GUS transient assay in the leaves of Nicotiana benthamiana revealed that the 162-bp (- 456~ - 295 bp) and 111-bp (- 294~ - 184 bp) fragments from the AtSCPL30 promoter could increase the transcriptional activity of mini35S up to 16- and 18-fold, respectively.
As a small constitutive strong promoter of plant origin, PD7 has the advantage of biosafety and reduces the probability of transgene silencing compared to the virus-derived CaMV35S promoter. PD7 would also be an alternative constitutive promoter to the CaMV35S promoter when multigene transformation was performed in the same vector, thereby avoiding the overuse of the CaMV35S promoter and allowing for the successful application of transgenic technology. And, the 162- and 111-bp fragments will also be very useful for synthetic promoter design based on their high enhancer activities.

Salinity and drought often affect plant growth and crop yields. Cloning and identification of salinity and drought stress inducible promoters is of great significance for their use in the genetic improvement of crop resistance. Previous studies showed that phosphatidylinositol synthase is involved in plant salinity and drought stress responses but its promoter has not been characterized by far. In the study, the promoter (pZmPIS, 1834 bp upstream region of the translation initiation site) was isolated from maize genome. To functionally validate the promoter, eight 5\' deletion fragments of pZmPIS in different lengths were fused to GUS to produce pZmPIS::GUS constructs and transformed into tobacco, namely PZ1-PZ8. The transcription activity and expression pattern obviously changed when the promoter was truncated. Previous studies have demonstrated that NaCl and PEG treatments are usually used to simulate salinity and drought treatments. The results showed that PZ1-PZ7 can respond well upon NaCl and PEG treatments, while PZ8 not. PZ7 (467 bp) displayed the highest transcription activity in all tissues of transgenic tobacco amongst 5\' deleted promoter fragments, which corresponds to about 20 and 50% of CaMV35S under normal and NaCl or PEG treatment, respectively. This implied that PZ7 is the core region of pZmPIS which confers high-level gene expression and NaCl or PEG inducible nature. The 113 bp segment between PZ7 and PZ8 (-467 to -355 bp) was considered as the key sequence for ZmPIS responding to NaCl or PEG treatment. GUS transient assay in tobacco leaves showed that this segment was sufficient for the NaCl or PEG stress response. Bioinformatic analysis revealed that the 113 bp sequence may contain new elements that are crucial for ZmPIS response to NaCl or PEG stress. These results promote our understanding on transcriptional regulation mechanism of ZmPIS and the characterized PZ7 promoter fragment would be an ideal candidate for the overexpression of drought and salinity responsive gene to improve crop resistance.