Transient gene expression of foreign genes in preheated protoplasts: stimulation of expression of transfected genes lacking heat shock elements.
Transfection of preheated petunia protoplasts with several biologically active DNA constructs resulted in a significantly higher gene expression than that observed in transfected unheated protoplasts. It was observed with supercoiled, linearized and single-stranded DNA structures that stimulation of transient gene expression in preheated protoplasts was neither dependent on the reporter gene nor on the regulatory elements used. Heat treatment at 42 degrees C also increased expression in protoplasts transfected with a plasmid bearing the tobacco mosaic virus (TMV) translational enhancer, omega. Northern blot analysis revealed that heat treatment of protoplasts before the transfection event greatly increased the amount of the newly synthesized transcripts. Preheating of protoplasts did not affect the transfection efficiency, namely the number of transfected cells in the population, nor the amount of DNA in transfected nuclei, as was inferred from histochemical staining and Southern blot analysis, respectively. The possible mechanism by which heat treatment stimulates transient gene expression of genes lacking obvious heat shock elements is offered. The relevance of the present findings to transient gene expression in plants in general and to viral gene expression in particular is discussed.
Zakai
N
N
Department of Biological Chemistry, Hebrew University of Jerusalem, Israel.
Ballas
N
N
Hershkovitz
M
M
Broido
S
S
Ram
R
R
Loyter
A
A
D002455:Cell Division; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D016384:Consensus Sequence; D005786:Gene Expression Regulation / Q000502:physiology*; D005966:Glucuronidase / Q000235:genetics; D006360:Heat-Shock Proteins / Q000235:genetics; D006358:Hot Temperature; D010944:Plants; D011523:Protoplasts / Q000378:metabolism*; D014162:Transfection / Q000379:methods*
D006360:Heat-Shock Proteins; D015500:Chloramphenicol O-Acetyltransferase; D005966:Glucuronidase