Dark quenchers for donor-acceptor energy transfer
The present invention provides a family of dark quenchers, termed Black Hole Quenchers (“BHQs”), that are efficient quenchers of excited state energy but which are themselves substantially non-fluorescent. Also provided are methods of using the BHQs, probes incorporating the BHQs and methods of using the probes.
1 - 87 . (canceled)
88 . A compound having the formula:
wherein
Ar 1 and Ar 2 are each independently a substituted or unsubstituted aryl group, where at least one of Ar 1 and Ar 2 is a substituted aryl;
X 1 is selected from the group consisting of OH, O-dimethoxytrityl, O-methoxytrityl, O-trityl
and an oxygen atom having an acid labile blocking group;
X 2 is a phosphoramidite; and
W is a linking group having from 3 to 100 backbone atoms selected from C, N, O, S, Si and P, said linking group being cyclic, acyclic, aromatic or a combination thereof.
89 . A compound having the formula:
wherein
Ar 1 and Ar 2 are each independently a substituted or unsubstituted aryl group, where at least one of Ar 1 and Ar 2 is a substituted aryl;
X 1 is selected from the group consisting of H, (C 1 -C 1-C12 )alkyl, aryl, heteroaryl, and protected or unprotected functional group;
X 2 is selected from the group consisting of a phosphorous coupling moiety, a pentafluorophenoxy moiety and a succinimidyl moiety; and
W is a linking group having from 3 to 100 backbone atoms selected from C, N, O, S, Si and P, said linking group being cyclic, acyclic, aromatic or a combination thereof.
90 . A compound having the formula:
wherein
Ar 1 and Ar 2 are each independently a substituted or unsubstituted aryl group, where at least one of Ar 1 and Ar 2 is a substituted aryl;
X 1 , is selected from the group consisting of H, (C 1 -C 12 )alkyl, aryl, heteroaryl and protected or unprotected functional group;
X 2 is a phosphoramidite; and
W is a linking group having from 3 to 100 backbone atoms selected from C, N, O, S, Si and P, said linking group being cyclic, acyclic, aromatic or a combination thereof.
91 . A compound having the formula:
wherein
Ar 1 and Ar 2 are each independently a substituted or unsubstituted aryl group;
X 1 is selected from the group consisting of H, (C 1 -C 12 )alkyl, aryl, heteroaryl, and protected or unprotected finctional group;
X 2 is a moiety reactive towards nucleophiles;
W is a linking group having from 3 to 100 backbone atoms selected from C, N, O, S, Si and P, said linking group being cyclic, acyclic, aromatic or a combination thereof; and
wherein one of Ar 1 and A 2 is directly or indirectly substituted with a substituted aryl group (A 3 ), where Ar 3 extends the resonance ability of the Ar 1 —N═N-A 2 aromatic system and thereby increases the wavelength absorbance maximum of the compound.
92 . A compound of claim 91 wherein Ar 1 is directly substituted with Ar 3 .
93 . A compound of claim 91 wherein Ar 1 is indirectly substituted with Ar 3 .
94 . A compound of claim 91 wherein A 2 is directly substituted with Ar 3 .
95 . A compound of claim 91 wherein Ar 2 is indirectly substituted with Ar 3 .
96 . A compound having the formula:
wherein
Ar 1 and A 2 are each independently a substituted or unsubstituted aryl group;
X 1 , is selected from the, group consisting of H, (C 1 -C 12 )alkyl, aryl, heteroaryl, and protected or unprotected functional group;
X 2 is a phosphoramidite;
W is a linking group having from 3 to 100 backbone atoms selected from C, N, O, S, Si and P, said linking group being cyclic, acyclic, aromatic or a combination thereof; and
wherein Ar 1 or Ar 2 is indirectly substituted with an aryl group through a group selected from —(C═C) n — and —(CR′CR′) n — where n is 0 to 5 and R′ is independently selected from hydrogen, (C 1 -C 8 )alkyl and heteroalkyl, unsubstituted aryl and heteroaryl, (unsubstituted aryl)-(C 1 -C 4 )alkyl, and (u nsubstituted aryl)oxy-(C 1 -C 4 )alkyl.
97 . A compound having the formula:
wherein
Ar 1 and Ar 2 are each independently a substituted or unsubstituted aryl group;
X 1 is selected from the group consisting of H, (C 1 -C 12 )alkyl, aryl, heteroaryl, and protected or unprotected functional group;
X 2 is a phosphoramidite;
W is a linking group having from 3 to 100 backbone atoms selected from C, N, O, S, Si and P, said linking group being cyclic, acyclic, aromatic or a combination thereof; and
Ar 1 or Ar 2 is indirectly substituted with an aryl group (Ar 3 ) through a double bond selected from carbon-carbon and nitrogen-nitrogen double bonds.
98 . A compound having the formula:
wherein
Ar 1 and A 2 are each independently a substituted or unsubstituted aryl group;
X 1 is selected from the group consisting of H, (C 1 -C 12 )alkyl, aryl, heteroaryl, and protected or unprotected functional group;
X 2 is a phosphoramidite;
W is a linking group having from 3 to 100 backbone atoms selected from C, N, O, S, Si and P, said linking group being cyclic, acyclic, aromatic or a combination thereof;
wherein one of Ar 1 and Ar 2 is directly or indirectly substituted with a substituted aryl group (Ar 3 ), where Ar 3 extends the resonance ability of the Ar 1 —N═N—Ar 2 aromatic system and thereby increases the wavelength absorbance maximum of the compound; and
at least one of Ar 1 , A 2 and Ar 3 is substituted with -halogen, —OR′, —OC(O)R′, —NR′R″, —SR′, —R′, —CN, —NO 2 , —CO 2 R′, —CONR′R″, —C(O)R′, —OC(O)NR′R″, —NR″C(O)R′, —NRC″C(O) 2 R′, —NR′—C(O)NR″R′″, —NH—C(NH 2 )NH, —NR′C(NH 2 )NH, —NH—C(NH 2 )═NR′, —S(O)R′, —S(O) 2 R′, —S(O) 2 NR′R″, —N3, —CH(Ph) 2 , perfluoro(C 1 -C 4 )alkoxy, and perfluoro(C 1 -C 4 )alkyl, in a number ranging from zero to the total number of open valences on the aromatic ring system; and where R′, R″ and R′″ are independently selected from hydrogen, (C 1 -C 8 )alkyl and heteroalkyl, unsubstituted aryl and heteroaryl, (unsubstituted aryl)-(C 1 -C 4 )alkyl, and (unsubstituted aryl)oxy-(C 1 -C 4 )alkyl.
99 . A compound of claim 88 wherein W is acylic.
100 . A compound of claim 88 wherein W comprises a cyclic group.
101 . A compound of claim 89 wherein W is a cyclic.
102 . A compound of claim 89 wherein W comprises a cyclic group.
103 . A compound of claim 90 wherein W is acyclic.
104 . A compound of claim 90 wherein W comprises a cyclic group.
105 . A compound of claim 91 wherein W is acyclic.
106 . A compound of claim 91 wherein W comprises a cyclic group.
107 . A compound of claim 92 wherein W is acyclic.
108 . A compound of claim 92 wherein W comprises a cyclic group.
109 . A compound of claim 93 wherein W is a cyclic.
110 . A compound of claim 93 wherein W comprises a cyclic group.
111 . A compound of claim 94 wherein W is acyclic.
112 . A compound of claim 94 wherein W comprises a cyclic group.
113 . A compound of claim 95 wherein W is acyclic.
114 . A compound of claim 95 wherein W comprises a cyclic group.
115 . A compound of claim 96 wherein W is acyclic.
116 . A compound of claim 96 wherein W comprises a cyclic group.
117 . A compound of claim 97 wherein W is cyclic.
118 . A compound of claim 97 wherein W comprises a cyclic group.
119 . A compound of claim 98 wherein W is acyclic.
120 . A compound of claim 98 wherein W comprises a cyclic group.
121 . A compound of claims 89 , 91 - 95 or 105 - 114 wherein X 2 is a phosphoramidite.