Author/Authors :
Seifi، Tahere نويسنده Department of Biology, School of Sciences, University of Isfahan , , Ghaedi، Kamran نويسنده , , Salamian، Ahmad نويسنده Department of Cell and Molecular Biology, Cell Science Research Center, Royan Institute for Animal Biotechnology, ACECR , , Tanhaei، Somayeh نويسنده , , Safari، Forouzan نويسنده Department of Biology, School of Sciences, University of Isfahan , , Hojati، Zohreh نويسنده , , Tavassoli، Manuchehr نويسنده Department of Biology, School of Sciences, University of Isfahan , , Baharvand، Hossein نويسنده , , NASR ESFAHANI، MOHAMMAD HOSSEIN نويسنده ,
Abstract :
Background: Recently, we have shown that peroxisomal protein expression was induced upon retinoic acid treatment in mouse embryonic stem cells during the process of neurogenesis. Thus, characterization of the respective promoter could elucidate the molecular aspects of transcriptional regulation of this gene.
Methods: Using the conventional software programs for promoter prediction, a putative promoter region was identified approximately 561 bp upstream of the peroxisomal protein coding sequence. In order to clone this region with a GC-content of 71.01%, a cocktail of ammonium sulfate buffer supplied with two additive components, betaine and dimethyl sulfoxide, and a high concentration of MgCl2 was used.
Results: The modulated polymerase chain reaction composition significantly improved the amplification of GC-rich DNA target sequences. Improved amplification of this region was due to reduction in the formation of secondary structures by the GC-rich region.
Conclusion: Therefore, this polymerase chain reaction composition could be generally used to facilitate the amplification of other GC-rich DNA sequences as verified by amplification of different GC rich regions.
