The role of green fluorescent protein (GFP) in transgenic plants to reduce gene silencing phenomena.
The green fluorescent protein (GFP) of jellyfish (Aequorea victoria) has significant advantages over other reporter genes, because expression can be detected in living cells without any substrates. Recently, epigenetic phenomena are important to consider in plant biotechnology experiments for elucidate unknown mechanism. Therefore, soybean immature cotyledons were generated embryogenesis cells and engineered with two different gene constructs (pHV and pHVS) using gene gun method. Both constructs contain a gene conferring resistance to hygromycin (hpt) as a selective marker and a modified glycinin (11S globulin) gene (V3-1) as a target. However, sGFP(S65T) as a reporter gene was used only in pHVS as a reporter gene for study the relation between using sGFP(S65T) and gene silencing phenomena. Fluorescence microscopic was used for screening after the selection of hygromycin, identified clearly the expression of sGFP(S65T) in the transformed soybean embryos bombarded with the pHVS construct. Protein analysis was used to detect gene expression overall seeds using SDS-PAGE. Percentage of gene down regulation was highly in pHV construct compared with pHVS. Thus, sGFP(S65T ) as a reporter gene in vector system may be play useful role for transgenic evaluation and avoid gene silencing in plants for the benefit of plant transformation system.
El-Shemy
Hany A
HA
Faculty of Agriculture Research Park (FARP) and Biochemistry Department, Faculty of Agriculture, Cairo University, Giza 12613, Egypt. helshemy@hotmail.com
Khalafalla
Mutasim M
MM
Ishimoto
Masao
M
D002934:Cinnamates / Q000378:metabolism; D003001:Cloning, Molecular; D018548:Cotyledon / Q000235:genetics / Q000502:physiology; D018506:Gene Expression Regulation, Plant / Q000502:physiology*; D020868:Gene Silencing / Q000502:physiology*; D017930:Genes, Reporter; D005916:Globulins / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D006921:Hygromycin B / Q000031:analogs & derivatives / Q000378:metabolism; D010940:Plant Proteins / Q000378:metabolism; D018520:Plant Shoots / Q000235:genetics / Q000502:physiology; D030821:Plants, Genetically Modified / Q000235:genetics / Q000502:physiology*; D030262:Soybean Proteins / Q000378:metabolism; D013025:Glycine max / Q000235:genetics / Q000502:physiology*
D002934:Cinnamates; D005916:Globulins; D010940:Plant Proteins; D030262:Soybean Proteins; D049452:Green Fluorescent Proteins; D006921:Hygromycin B; C026273:hygromycin A; C011474:glycinin