Abstract:
Even though it is the staple food of more than half the world’s population, rice is not
considered a rich source of protein. One of the essential amino acids, lysine is low in rice
seed. Hence, this study was aimed at increasing both the lysine and total protein content
in rice seeds by introducing the pollen-specific lysine-rich protein encoding gene (SBgLR)
from potato (Solanum tuberosum) into indica rice (Oryza sativa L.) seed under the control
of the rice seed-specific globulin promoter. Total Ribo Nucleic Acid (RNA) from potato
pollen grains was extracted and the produced CDNA and SBgLR gene was amplified by
Polymerase Chain Reaction (PCR) using SBgLR gene-specific primers. The isolated rice
genomic Deoxy Ribonucleic Acid (DNA) from rice variety Bg 94-1 was subjected to PCR to
amplify the promoter sequence of the globulin gene. The amplified promoter region was
cloned into pGEM®-T Easy vector, and then into pCAMBIA1391Z vector. Recombinants
were selected and sequenced. The SBgLR gene containing recombinant vector (pCR®2.1-
TOPO-SBgLR), previously cloned in our laboratory was restriction digested with BamH1
and EcoR1 enzymes, and cloned into the corresponding sites of pCAMBIA1391Z-Glb
vector construct. Electro competent cells of Agrobacterium strain GV3101 was transformed
with the recombinant construct (pCAMBIA1391Z-Glb-SBgLR). Bg 94-1 rice seed derived
calli, and active rice embryos were used for transformation. The embryo transformation
method proved less time consuming and more effective in producing transgenic rice plants.
PCR analysis of regenerated transformed plants indicated the presence of the SBgLR
gene.
