CH391L/S14/Non-canonical Nucleotides

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Non-canonical nucleobases are small heterocyclic molecules that function similar to the natural nucleobases by pairing through hydrogen bounding, hydrophobic interactions, or other means. These base-pairing capabilities are essential in functioning like the natural base pairs. Non-canonical nucleobases are used, or proposed to use for, for a variety of purposes, including improving the theoretical efficiency of DNA computing, researching the evolution of life, and containing synthetic life. Several research groups are working on non-canonical nucleobases, and utilizing them in different ways. Despite advances, there are several significant obstacles to implementing non-canonical nucleobases in vivo.

Contents

History and Examples

A collection of several non-canonical base pairs that have been developed[1]
It was proposed in 1962 that a third possible DNA base-pair might occur in nature, the iso-C/Iso-G pairing. This pairing was unobserved until 1989, when the Benner group first demonstrated that the iso-C/iso-G pair could be enzymatically incorporated from a template containing iso-C, using an E. coli DNA polymerase fragment[2]. Additionally, it was shown that T7 RNA polymerase was capable of transcribing iso-C/iso-G as a third set of base pairs with a template DNA sequencing containing iso-C and iso-G.

Since the original paper demonstrating the propagation of non-canonical bases in DNA and RNA, several labs have published other base pairs that are capable of preservation in DNA and RNA.

xDNA

xDNA is a is an expanded DNA set with the four nature nucleobases pairing with four size expanded nucleobases. The four size expanded nucleobases are natural nucleobases with an additional benzene ring, so there exists adenine
The base-pairing patterns of xDNA, showing the expanded bases pairing with natural bases.
(A), thymine (T), cytosine (C), and guanine (G), and there exists xA, xT, xC, and xG. In xDNA, a natural base pairs with an extended base, so A will pair with xT, and G will pair with xC. This pairing preserve the natural hydrogen bonding formations, but widens the helix throughout the entire DNA molecule by 2.4 Angstroms[3]. xDNA is more thermal stable then natural DNA, and can base pair with natural RNA and DNA of up to four bases in length quite well, but as then length approaches eight base pairs or longer, xDNA does not hybridize with DNA or RNA. This would make xDNA orthogonal to DNA, while still preserving some of the features, and would greatly expand the code by doubling the available bases. Additionally, xDNA regions of up to four base pairs have been preserved in vivo in plasmids carried by E. coli [3].

Hirao Bases

Ichiro Hirao at RIKEN and TagCyx biotechnologies have developed a set of base pairs build around Ds and a variety of pairing options, initially Pa, but subsequently published base pairing with Pn and Px. The Hirao bases are efficiently propagated in normal PCR conditions[4].
The Hirao base pairs, Pn and Px are capable of replication using PCR without modification, with DsTPs and PxTPs
. Hirao used his bases in aptamer selections, and pulled out aptamers with 100 fold improvement affinity for their targets compared to the previously published aptamers for the target molecules(VEGF-165 and interferon-gamma)[5].


X-DAP F-Za S-Pa Ds-Px Others Diaminopurine xDNA


Uses Aptamers -fluorescent base pairs

Challenges

Future

Steven Benner is working on creating

iGEM

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  1. Error fetching PMID 19318205: [Kool2009]
  2. Error fetching PMID 1688644: [Benner1990]
  3. Error fetching PMID 21981660: [Kruger2012]
  4. Error fetching PMID 18776457: [Hirao2008]
  5. Error fetching PMID 23563318: [Hirao2013]
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