Novel Strategies to Optimize the Amplification of Single-Stranded DNA - Archive ouverte HAL Accéder directement au contenu
Article Dans Une Revue Frontiers in Bioengineering and Biotechnology Année : 2020

Novel Strategies to Optimize the Amplification of Single-Stranded DNA

Résumé

The generation of single stranded DNA plays a key role in in vitro selection of DNA aptamers and in other molecular techniques such as DNA sequencing and microarrays. Here we describe three novel methodologies for ssDNA production and amplification. Furthermore, we describe some previously unnoticed aspects of random DNA amplification. Our results showed that in asymmetric PCR the addition of a high melting temperature reverse primer blocked at its 3' end by a dideoxy nucleotide drives the reaction further toward ssDNA production. We demonstrated also that incorporation of internally inverted nucleotide/(s) in one primer can be used as a new method of polymerization termination. Using such modified primer, the PCR product includes two complementary DNA strands having different lengths and separable from one another by denaturing gel electrophoresis. In addition, we showed that nicking enzymes can be used to cleave the undesirable strand allowing the isolation of the target ssDNA strand.
Fichier principal
Vignette du fichier
fbioe-08-00401.pdf (7.21 Mo) Télécharger le fichier
Origine : Publication financée par une institution

Dates et versions

hal-03590600 , version 1 (28-02-2022)

Licence

Paternité

Identifiants

Citer

Atef Nehdi, Nosaibah Samman, Vanessa Aguilar-Sánchez, Azer Farah, Emre Yurdusev, et al.. Novel Strategies to Optimize the Amplification of Single-Stranded DNA. Frontiers in Bioengineering and Biotechnology, 2020, 8, pp.401. ⟨10.3389/fbioe.2020.00401⟩. ⟨hal-03590600⟩

Collections

RIIP INRS-IAF
45 Consultations
6 Téléchargements

Altmetric

Partager

Gmail Facebook X LinkedIn More