Upgrading SELEX technology by using lambda exonuclease digestion for single-stranded DNA generation.
The generation of single-stranded DNA (ssDNA) molecules plays a key role in the SELEX (Systematic Evolution of Ligands by EXponential enrichment) combinatorial chemistry process and numerous molecular biology techniques and applications, such as DNA sequencing, single-nucleotide polymorphism (SNP) analysis, DNA chips, DNA single-strand conformation polymorphism (SSCP) analysis and many other techniques. The purity and yield of ssDNA can affect the success of each application. This study compares the two ssDNA production methods, the strand separation by streptavidin-coated magnetic beads and alkaline denaturation and the lambda exonuclease digestion, in regard to the purity of generated ssDNA and the efficiency. Here, we demonstrate the considerable benefits of ssDNA production by lambda exonuclease digestion for in vitro selection of DNA aptamers. We believe that the generation of ssDNA aptamers using this method will greatly improve the success rate of SELEX experiments concerning the recovery of target-specific aptamers.
Avci-Adali
Meltem
M
Department of Thoracic, Cardiac, and Vascular Surgery, University Hospital Tuebingen, Calwerstr. 7/1, Tuebingen 72076, Germany. hans-peter.wendel@med.uni-tuebingen.de
Paul
Angel
A
Wilhelm
Nadj
N
Ziemer
Gerhard
G
Wendel
Hans Peter
HP
D019921:Biotinylation / Q000187:drug effects; D015153:Blotting, Western; D004277:DNA, Single-Stranded / Q000096:biosynthesis*; D005090:Exodeoxyribonucleases / Q000378:metabolism*; D008280:Magnetics; D008863:Microspheres; D009691:Nucleic Acid Denaturation / Q000187:drug effects; D010766:Phosphorylation / Q000187:drug effects; D052156:SELEX Aptamer Technique / Q000379:methods*; D012972:Sodium Hydroxide / Q000494:pharmacology; D019809:Streptavidin / Q000378:metabolism; D013997:Time Factors
D004277:DNA, Single-Stranded; D012972:Sodium Hydroxide; D019809:Streptavidin; D005090:Exodeoxyribonucleases