Abstract
From a library of nucleic acid molecules, which are randomized in parts of their sequence, unique sequence variants can be selected for specific properties. The planning of such an in vitro selection experiment requires some consideration regarding how much DNA template or RNA transcript should be used initially. The amount applied depends on the number of randomized nucleotides and on the expectations of how often each conceivable and unique sequence combination should be represented in the experimental pool. We display graphs describing the probability for the representation of unique nucleic acid molecules in a randomized pool as a function of the mean representation k, defined by the ratio of sampled nucleic acid molecules to conceivable sequence combinations and we summarize the amounts required to represent unique sequences with 99% likelihood. The probability of representation, P = 1-e-k, can be applied also to 'sub-saturated' pools (k < 1) of nucleic acids with long randomized domains, where it is impossible to provide sufficient material for full sequence representation.
Full Text
The Full Text of this article is available as a PDF (34.5 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Beaudry A. A., Joyce G. F. Directed evolution of an RNA enzyme. Science. 1992 Jul 31;257(5070):635–641. doi: 10.1126/science.1496376. [DOI] [PubMed] [Google Scholar]
- Berzal-Herranz A., Joseph S., Burke J. M. In vitro selection of active hairpin ribozymes by sequential RNA-catalyzed cleavage and ligation reactions. Genes Dev. 1992 Jan;6(1):129–134. doi: 10.1101/gad.6.1.129. [DOI] [PubMed] [Google Scholar]
- Gold L., Polisky B., Uhlenbeck O., Yarus M. Diversity of oligonucleotide functions. Annu Rev Biochem. 1995;64:763–797. doi: 10.1146/annurev.bi.64.070195.003555. [DOI] [PubMed] [Google Scholar]
- Irvine D., Tuerk C., Gold L. SELEXION. Systematic evolution of ligands by exponential enrichment with integrated optimization by non-linear analysis. J Mol Biol. 1991 Dec 5;222(3):739–761. doi: 10.1016/0022-2836(91)90509-5. [DOI] [PubMed] [Google Scholar]
- Kumar P. K., Ellington A. D. Artificial evolution and natural ribozymes. FASEB J. 1995 Sep;9(12):1183–1195. doi: 10.1096/fasebj.9.12.7672511. [DOI] [PubMed] [Google Scholar]
- Robertson D. L., Joyce G. F. Selection in vitro of an RNA enzyme that specifically cleaves single-stranded DNA. Nature. 1990 Mar 29;344(6265):467–468. doi: 10.1038/344467a0. [DOI] [PubMed] [Google Scholar]
- Tuerk C., Gold L. Systematic evolution of ligands by exponential enrichment: RNA ligands to bacteriophage T4 DNA polymerase. Science. 1990 Aug 3;249(4968):505–510. doi: 10.1126/science.2200121. [DOI] [PubMed] [Google Scholar]