Assay of insulator enhancer-blocking activity with the use of transient transfection.
We used a transient transfection of cultured cells with linearized plasmids to analyze the enhancer-blocking activity of potential insulators including the standard cHS4 chicken beta-globin insulator and several DNA fragments selected from the human genome sequence. About 60-80% of the potential insulators do reveal the enhancer-blocking activity when probed by the transient transfection assay. The activity of different sequences is characterized by certain tissue specificity and by dependence on the orientation of the fragments relative to the promoter. Thus, the transfection model may be used for quantitative analysis of the enhancer-blocking activity of the potential insulators.
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, 117997, Russia. akser@ibch.ru.
N A
NA
Smirnov
D A
DA
Didych
S B
SB
Akopov
L G
LG
Nikolaev
E D
ED
Sverdlov
D000818:Animals; D001681:Biological Assay; D016466:CHO Cells; D002645:Chickens; D003412:Cricetulus; D004247:DNA / Q000235:genetics; D004742:Enhancer Elements, Genetic; D015894:Genome, Human; D006367:HeLa Cells; D056945:Hep G2 Cells; D006801:Humans; D038101:Insulator Elements; D010957:Plasmids; D011401:Promoter Regions, Genetic; D014162:Transfection; D055544:beta-Globins / Q000235:genetics / Q000378:metabolism
D055544:beta-Globins; D004247:DNA