title: Novel cyanine-AMP conjugates for efficient 5' RNA fluorescent labeling by one-step transcription and replacement of [gamma-32P]ATP in RNA structural investigation.
abstract: Two novel fluorescent cyanine-AMP conjugates, F550/570 and F650/670, have been synthesized to serve as transcription initiators under the T7 phi2.5 promoter. Efficient fluorophore labeling of 5' RNA is achieved in a single transcription step by including F550/570 and F650/670 in the transcription solution. The current work makes fluorescently labeled RNA readily available for broad applications in biochemistry, molecular biology, structural biology and biomedicine. In particular, site-specifically fluorophore-labeled large RNAs prepared by the current method may be used to investigate RNA structure, folding and mechanism by various fluorescence techniques. In addition, F550/570 and F650/670 may replace [gamma-32P]ATP to prepare 5' labeled RNA for RNA structural and functional investigation, thereby eliminating the need for the unstable and radio-hazardous [gamma-32P]ATP.
authors.affiliation: Department of Chemistry and Biochemistry, University of Southern Mississippi Hattiesburg MS 39406-5043, USA.
authors.forename: Na
authors.initials: N
authors.lastname: Li
authors.forename: Changjun
authors.initials: C
authors.lastname: Yu
authors.forename: Faqing
authors.initials: F
authors.lastname: Huang
mesh_terms: D000249:Adenosine Monophosphate / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry*; D000255:Adenosine Triphosphate / Q000737:chemistry; D001483:Base Sequence; D005456:Fluorescent Dyes / Q000138:chemical synthesis / Q000737:chemistry*; D007211:Indoles / Q000138:chemical synthesis / Q000737:chemistry*; D008969:Molecular Sequence Data; D010761:Phosphorus Radioisotopes; D012313:RNA / Q000096:biosynthesis / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet; D014158:Transcription, Genetic
chemical_list: C497832:F550-570 compound; C497833:F650-670 compound; D005456:Fluorescent Dyes; D007211:Indoles; D010761:Phosphorus Radioisotopes; D000249:Adenosine Monophosphate; D012313:RNA; D000255:Adenosine Triphosphate
references.citation: RNA. 1999 Oct;5(10):1348-56
references.pmid: 10573125
references.citation: Biochemistry. 2005 Mar 22;44(11):4582-90
references.pmid: 15766289
references.citation: Methods. 2001 Mar;23(3):240-54
references.pmid: 11243837
references.citation: EMBO J. 2001 Apr 17;20(8):2051-61
references.pmid: 11296237
references.citation: Methods. 2001 Sep;25(1):19-30
references.pmid: 11558994
references.citation: Proc Natl Acad Sci U S A. 2002 Apr 2;99(7):4284-9
references.pmid: 11929999
references.citation: Biopolymers. 2001-2002;61(3):159-79
references.pmid: 11987179
references.citation: Biopolymers. 2001-2002;61(3):224-42
references.pmid: 11987183
references.citation: RNA. 2002 Oct;8(10):1242-52
references.pmid: 12403463
references.citation: Chem Biol. 2002 Nov;9(11):1227-36
references.pmid: 12445773
references.citation: Nucleic Acids Res. 2003 Feb 1;31(3):e8
references.pmid: 12560511
references.citation: Curr Opin Struct Biol. 2003 Feb;13(1):88-97
references.pmid: 12581665
references.citation: RNA. 2003 Dec;9(12):1562-70
references.pmid: 14624011
references.citation: Proc Natl Acad Sci U S A. 2004 Jan 13;101(2):534-9
references.pmid: 14704266
references.citation: Methods Mol Biol. 2004;252:77-108
references.pmid: 15017044
references.citation: Nucleic Acids Res. 2004;32(9):2707-15
references.pmid: 15148358
references.citation: Proc Natl Acad Sci U S A. 2004 Jul 6;101(27):10066-71
references.pmid: 15218105
references.citation: Cytometry. 1990;11(3):418-30
references.pmid: 2340776
references.citation: Science. 1991 Jan 25;251(4992):401-7
references.pmid: 1989074
references.citation: Biochemistry. 1991 May 14;30(19):4821-30
references.pmid: 2029524
references.citation: Science. 1994 Nov 4;266(5186):785-9
references.pmid: 7973630
references.citation: Bioconjug Chem. 1993 Mar-Apr;4(2):105-11
references.pmid: 7873641
references.citation: Biochemistry. 1997 Apr 22;36(16):4718-30
references.pmid: 9125492
references.citation: Science. 1998 Mar 20;279(5358):1940-3
references.pmid: 9506944
references.citation: EMBO J. 1998 Apr 15;17(8):2378-91
references.pmid: 9545249
references.citation: Methods. 1999 May;18(1):60-70
references.pmid: 10208817
references.citation: Methods Enzymol. 2000;317:409-40
references.pmid: 10829293