Probes labeled with energy transfer coupled dyes
First Claim
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1. A fluorescent label comprising:
- at least two fluorophores bonded to a polymeric backbone at specific locations thereon to form a chemically pure label wherein at least one fluorophore is bonded to a terminal monomeric unit of said polymeric backbone and the other fluorophore(s) is bonded to a non-terminal monomeric unit of said polymeric backbone and said terminally bonded fluorophore is in an energy transfer relationship with said other fluorophore(s), said fluorophores comprising a donor fluorophore and an acceptor fluorophore of which said donor fluorophore absorbs light in the wavelength range of 350 to 800 nm and said acceptor emits light in the wavelength range of 450 to 1000 nm.
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Abstract
Fluorescent labels having at least one donor and at least one acceptor fluorophore bonded to a polymeric backbone in energy transfer relationship, as well as methods for their use, are provided. Of particular interest are the subject labels wherein the polymeric backbone is a nucleic acid and the donor fluorophore is bonded to the 5'"'"' terminus of said nucleic acid. Such labels find use as primers in applications involving nucleic acid chain extension, such as sequencing, PCR and the like.
179 Citations
27 Claims
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1. A fluorescent label comprising:
at least two fluorophores bonded to a polymeric backbone at specific locations thereon to form a chemically pure label wherein at least one fluorophore is bonded to a terminal monomeric unit of said polymeric backbone and the other fluorophore(s) is bonded to a non-terminal monomeric unit of said polymeric backbone and said terminally bonded fluorophore is in an energy transfer relationship with said other fluorophore(s), said fluorophores comprising a donor fluorophore and an acceptor fluorophore of which said donor fluorophore absorbs light in the wavelength range of 350 to 800 nm and said acceptor emits light in the wavelength range of 450 to 1000 nm. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8)
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9. A fluorescent label comprising:
at least two fluorophores comprising a donor fluorophore and an acceptor fluorophore bonded to an oligonucleotide at specific locations thereon to form a chemically pure label, wherein one of said fluorophores is bonded to the 5'"'"' terminus of said oligonucleotide and the other of said fluorophores is bonded to a modified nucleotide from 2 to 20 nucleotides 3'"'"' of said 5'"'"' bonded fluorophore and said 5'"'"' bonded fluorophore is in energy transfer relationship with said other fluorophore, and wherein said donor fluorophore absorbs light in the wavelength range of 350 to 800 nm and said acceptor fluorophore emits light in the wavelength range of 450 to 1000 nm. - View Dependent Claims (10, 11, 12, 13, 14, 15)
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16. A fluorescent label comprising:
donor and acceptor fluorophores having a difference in emission maxima that does not exceed 100 nm bonded to an oligonucleotide at specific locations thereon to form a chemically pure label, wherein one of said donor and acceptor fluorophores is bonded to the 5'"'"' terminus of said oligonucleotide and the other of said donor and acceptor fluorophores is bonded to a modified nucleotide from 2 to 20 nucleotides 3'"'"' of said 5'"'"' bonded fluorophore. - View Dependent Claims (17, 18, 19, 20, 21)
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22. A fluorescent label comprising:
a donor fluorophore and an acceptor fluorophore having a difference in emission maxima that does not exceed 100 nm bonded to an oligonucleotide at specific locations thereon to form a chemically pure label, wherein one of said donor and acceptor fluorophores is bonded to the sugar group of the 5'"'"' nucleotide of said oligonucleotide and the other of said donor and acceptor fluorophores is bonded to a modified nucleotide from 2 to 15 nucleotides 3'"'"' of 5'"'"' bonded fluorophore, wherein said modified nucleotide is Amino Modifier C6 dT. - View Dependent Claims (23)
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24. A fluorescent label having a donor fluorophore and an acceptor fluorophore bonded to an oligonucleotide at specific locations thereon to form a chemically pure label, wherein said donor fluorophore is bonded to the sugar group of the 5'"'"' nucleotide of said oligonucleotide and said acceptor fluorophore is bonded to a modified nucleotide 3'"'"' of said 5'"'"' bonded fluorophore, said label described by the formula:
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space="preserve" listing-type="equation">D--N--Awherein; D is FAM; N is the number of bases between said donor fluorophore and said acceptor fluorophore and ranges from 2 to 14; and A is FAM.
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25. A fluorescent label having a donor fluorophore and an acceptor fluorophore bonded to an oligonucleotide at specific locations thereon to form a chemically pure label, wherein said donor fluorophore is bonded to the sugar group of the 5'"'"' nucleotide of said oligonucleotide and said acceptor fluorophore is bonded to a modified nucleotide 3'"'"' of said 5'"'"' bonded fluorophore, said label described by the formula:
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space="preserve" listing-type="equation">D--N--Awherein; D is FAM; N is the number of bases between said donor fluorophore and said acceptor fluorophore and ranges from 2 to 14; and A is JOE.
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26. A fluorescent label having a donor fluorophore and an acceptor fluorophore bonded to an oligonucleotide at specific locations thereon to form a chemically pure label, wherein said donor fluorophore is bonded to the sugar group of the 5'"'"' nucleotide of said oligonucleotide and acceptor fluorophore is bonded to a modified nucleotide 3'"'"' of said 5'"'"' bonded fluorophore, said label described by the formula:
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space="preserve" listing-type="equation">D--N--Awherein; D is FAM; N is the number of bases between said donor fluorophore and said acceptor fluorophore and ranges from 2 to 14; and A is TAMRA.
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27. A fluorescent label having a donor fluorophore and an acceptor fluorophore bonded to an oligonucleotide at specific locations thereon to form a chemically pure label, wherein said donor fluorophore is bonded to the sugar group of the 5'"'"' nucleotide of said oligonucleotide and said acceptor fluorophore is bonded to a modified nucleotide 3'"'"' of said 5'"'"' bonded fluorophore, said label described by the formula:
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space="preserve" listing-type="equation">D--N--Awherein; D is FAM; N is the number of bases between said donor fluorophore and said acceptor fluorophore and ranges from 2 to 14; and A is ROX.
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Specification