Temporal and spatial expression analysis of gamma kafirin promoter from Sorghum (Sorghum bicolor L. moench) var. M 35-1.
Different cis acting elements of gamma kafirin gene from Sorghum bicolor var. M 35-1 were amplified and cloned using different combination of the primers. The amplified promoter was replaced with CaMV35S promoter of vector pCMBIA-1304 and resultant vector contained beta-glucuronidase (gus) gene under the control of amplified gamma-kafirin promoter. The resulting fusants were then transformed in to different explants of sorghum via particle bombardment. The regulation of uid gene expression was analyzed to find out the minimum required 5' regulatory sequence and cis acting elements for the efficient expression. However no gus expression was detected in leaves of micropropagated plants, scutellum and calli at any stage of growth. The expression of gus, with pKaf gus-P4 gene construct, was detected in immature embryos and endosperm 20 days after pollination (DAP). The result suggest that at least three motifs (two GCN4 and one prolamin box) besides TATA and CATC boxes are required for the efficient expression of the kafirin gene of sorghum. The study shows that PCR based isolation of different motifs and regions can be used as an alternate to deletion analysis for observing the role of various motifs and their importance in the gene expression and regulation.
Mishra
Avinash
A
Molecular Biology and Biotechnology, Central Salt and Marine Chemicals Research Institute, G. B. Marg, Bhavnagar, Gujarat, 364 002, India. avinash@csmcri.org
Tomar
Akash
A
Bansal
Sangita
S
Khanna
V K
VK
Garg
G K
GK
D001483:Base Sequence; D002874:Chromosome Mapping; D017931:DNA Primers / Q000378:metabolism; D018744:DNA, Plant / Q000235:genetics / Q000302:isolation & purification; D018506:Gene Expression Regulation, Plant; D005822:Genetic Vectors / Q000235:genetics; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000235:genetics*; D055315:Prolamins; D011401:Promoter Regions, Genetic / Q000235:genetics*; D017422:Sequence Analysis, DNA; D045868:Sorghum / Q000235:genetics*; D013997:Time Factors
D017931:DNA Primers; D018744:DNA, Plant; D010940:Plant Proteins; D055315:Prolamins; C080615:kafirin protein, Sorghum bicolor