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Enzymatic synthesis of DNA strands containing α-L-LNA (α-L-configured locked nucleic acid) thymine nucleotides.

Højland T, Veedu RN, Vester B, Wengel J - Artif DNA PNA XNA (2012 Jan-Mar)

Bottom Line: It was found that the 5'-triphosphate of α-L-LNA is a substrate for the DNA polymerases KOD, 9°N(m), Phusion and HIV RT.Three dispersed α-L-LNA thymine nucleotides can be incorporated into DNA strands by all four polymerases, but they were unable to perform consecutive incorporations of α-L-LNA nucleotides.In addition it was found that primer extension can be achieved using templates containing one α-L-LNA nucleotide.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics, Chemistry and Pharmacy, Nucleic Acid Center, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark.

ABSTRACT
We describe the first enzymatic incorporation of an α-L-LNA nucleotide into an oligonucleotide. It was found that the 5'-triphosphate of α-L-LNA is a substrate for the DNA polymerases KOD, 9°N(m), Phusion and HIV RT. Three dispersed α-L-LNA thymine nucleotides can be incorporated into DNA strands by all four polymerases, but they were unable to perform consecutive incorporations of α-L-LNA nucleotides. In addition it was found that primer extension can be achieved using templates containing one α-L-LNA nucleotide.

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Scheme 1. Chemical synthesis of α-L-LNA TTP. (i) Cl2CHCOOH, Et3SiH, CH2Cl2 (100%); (ii) (1) (MeO)3PO, proton sponge, POCl3, -10°C → -5°C; (2) tri-n-butylamine, tributylammonium pyrophosphate, dimethylformamide, -5°C; (3) triethylammonium bicarbonate (4%).
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S1: Scheme 1. Chemical synthesis of α-L-LNA TTP. (i) Cl2CHCOOH, Et3SiH, CH2Cl2 (100%); (ii) (1) (MeO)3PO, proton sponge, POCl3, -10°C → -5°C; (2) tri-n-butylamine, tributylammonium pyrophosphate, dimethylformamide, -5°C; (3) triethylammonium bicarbonate (4%).


Enzymatic synthesis of DNA strands containing α-L-LNA (α-L-configured locked nucleic acid) thymine nucleotides.

Højland T, Veedu RN, Vester B, Wengel J - Artif DNA PNA XNA (2012 Jan-Mar)

Scheme 1. Chemical synthesis of α-L-LNA TTP. (i) Cl2CHCOOH, Et3SiH, CH2Cl2 (100%); (ii) (1) (MeO)3PO, proton sponge, POCl3, -10°C → -5°C; (2) tri-n-butylamine, tributylammonium pyrophosphate, dimethylformamide, -5°C; (3) triethylammonium bicarbonate (4%).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3368812&req=5

S1: Scheme 1. Chemical synthesis of α-L-LNA TTP. (i) Cl2CHCOOH, Et3SiH, CH2Cl2 (100%); (ii) (1) (MeO)3PO, proton sponge, POCl3, -10°C → -5°C; (2) tri-n-butylamine, tributylammonium pyrophosphate, dimethylformamide, -5°C; (3) triethylammonium bicarbonate (4%).
Bottom Line: It was found that the 5'-triphosphate of α-L-LNA is a substrate for the DNA polymerases KOD, 9°N(m), Phusion and HIV RT.Three dispersed α-L-LNA thymine nucleotides can be incorporated into DNA strands by all four polymerases, but they were unable to perform consecutive incorporations of α-L-LNA nucleotides.In addition it was found that primer extension can be achieved using templates containing one α-L-LNA nucleotide.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics, Chemistry and Pharmacy, Nucleic Acid Center, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark.

ABSTRACT
We describe the first enzymatic incorporation of an α-L-LNA nucleotide into an oligonucleotide. It was found that the 5'-triphosphate of α-L-LNA is a substrate for the DNA polymerases KOD, 9°N(m), Phusion and HIV RT. Three dispersed α-L-LNA thymine nucleotides can be incorporated into DNA strands by all four polymerases, but they were unable to perform consecutive incorporations of α-L-LNA nucleotides. In addition it was found that primer extension can be achieved using templates containing one α-L-LNA nucleotide.

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