A phosphoramidite RO sub 2 sub PNR sub 2 sub is a monoamide of a phosphite diester. The key feature of phosphoramidites is their markedly high reactivity towards nucleophile s catalyzed by weak acids e.c ., triethylammonium chloride or 1 H tetrazole . In these reactions, the incoming nucleophile replaces the NR sub 2 sub moiety. Applications Nucleoside phosphoramidites main Nucleoside phosphoramidite Phosphoramidites derived from protected nucleosides are referred to as nucleoside phosphoramidite s and are widely used in Oligonucleotide synthesis chemical synthesis of DNA , RNA, and other nucleic acids and their analogs. As ligands Certain phosphoramidites are also used as monodentate chiral ligand s in asymmetric synthesis. ref cite DOI 10.1002 anie.200904948 ref A large group of such ligands is derived from the chiral diol BINOL and can be synthesis by reaction of BINOL with phosphorus trichloride to the chlorophosphite and then reaction with simple secondary amine s. ref cite DOI 10.1016 0957 4166 94 80032 4 ref This type of ligand was first used in 1996 in an asymmetric copper catalysed addition of dialkylzincs to enones ref cite DOI 10.1002 anie.199623741 ref ref cite DOI 10.1002 anie.199726201 ref See also Phosphoramidate References Reflist organophosphorus Category Phosphoramidites pl Amidofosforyny ... more details
, the use of the appropriate protecting strategy allows one to convert that to the respective phosphoramidite ... of an oligonucleotide chain using phosphoramidite strategy, the nucleoside analog have to possess ... on silica gel . To warrant the stability of the phosphoramidite moiety, it is advisable ... the entire course of the separation. The purity of a phosphoramidite may be assessed by sup 31 sup P NMR spectroscopy . As the P III atom is chiral, in a nucleoside phosphoramidite it displays two peaks ... H phosphonate impurities display peaks at 8 and 10 ppm. Chemical properties of phosphoramidite moiety ... important feature of phosphoramidites is their ability to undergo the phosphoramidite coupling reaction ... 1H tetrazole as activator for 2 O TBDMS phosphoramidite building blocks in RNA synthesis ... useful intermediates in oligonucleotide synthesis . Stereochemically, the phosphoramidite coupling ... oxidized diastereomeric 5 dimethoxytrityl thymidine 3 O O 2 cyanoethyl N,N diisopropyl phosphoramidite ... improved by using 3 O N , N diisopropyl phosphoramidite derivatives of nucleosides nucleoside ... 2008 08 16 accessdate 2009 05 12 ref and most common nucleoside phosphoramidite building blocks are briefly ... N isobutyryl protected deoxyadenosine phosphoramidite year 1997 author McMinn, D. journal Tetrahedron ... oligonucleotide synthesis. XVIII use of cyanoethyl N,N dialkylamino N morpholino phosphoramidite ... pmid 6547529 issue 11 pmc 318857 ref Once a phosphoramidite has been coupled to the solid support ... 66 issue 5 pages 1798 1804 doi 10.1021 jo001591e title Phosphoramidite Coupling to Oligonucleotides ... by the phosphoramidite approach. Tetrahedron 1992 , 48 , 2223 2311. Beaucage, S. L. Iyer, R. P. The functionalization of oligonucleotides via phosphoramidite derivatives. Tetrahedron 1993 , 49 , 1925 1963. Beaucage, S. L. Iyer, R. P. The synthesis of modified oligonucleotides by the phosphoramidite ... synthesis. Phosphoramidite approach. Methods in Molecular Biology Totowa, NJ, United States ... more details
A phosphoramidate is a phosphate that has an NR sub 2 sub instead of an OH group. The structure of phosphoramidic acid phosphoramidate , HO sub 2 sub PONH sub 2 sub , is present in PubChem ref http pubchem.ncbi.nlm.nih.gov summary summary.cgi?cid 211207 ref . br A phosphorodiamidate is a phosphate that has two of its OH groups substituted by NR sub 2 sub groups. Examples An example of a natural phosphoramidate is phosphocreatine . An example of a phosphorodiamidate is Morpholino which is used in molecular biology . See also Phosphoramidite References references organophosphorus Category Phosphoramidates ... more details
. In addition to the conventional set of nucleoside phosphoramidite s, the synthesis also requires a solid support derivatized with a quencher and a phosphoramidite building block designed for the attachment ... more details
Refimprove article date June 2010 Fluorescent labelling is the process of covalently attaching a fluorophore to another molecule, such as a protein or nucleic acid . This is generally accomplished using a reactive derivative of the fluorophore that selectively binds to a functional group contained in the target molecule. The most commonly labelled molecules are antibodies, proteins, amino acids and peptides which are then used as specific probes for detection of a particular target. ref http pharmaxchange.info press 2011 01 fluorescent labeling of biomolecules with organic probes Presentation on Fluorescent labelling of biomolecules with organic probes PharmaXChange.info ref Detection Fluorescent labels are generally used for detection of a protein or other labeled molecule via a fluorescence microscope , flow cytometer or some other fluorescence reading instrument. These can be useful in localization of a target within a cell, flow cytometry FACS analysis, western blot assays, and other immunoanalytical methods. Labelling techniques Fluorescent labelling is accomplished using a chemically reactive derivative of a fluorophore. Common reactive groups include Isothiocyanate derivatives such as Fluorescein isothiocyanate FITC and TRITC derivatives of fluorescein and rhodamine are reactive towards primary amines to form a thioureido linkage between the compound of interest and the dye. Succinimidyl esters such as NHS fluorescein are reactive towards amino groups to form an amido bond. Maleimide activated fluorophores such as fluorescein 5 maleimide readily react with sulfhydryl groups. The sulfhydryl group adds to the double bond of the maleimide. In oligonucleotide synthesis , several phosphoramidite reagents containing protected fluorescein and other fluorophores, e.g. fluorescein amidite 6 FAM phosphoramidite 2 , ref Brush, C. K. Fluorescein Labelled Phosphoramidites. US Patent 5,583,236. http patft.uspto.gov netacgi nph Parser?Sect1 PTO2&Sect2 HITOFF&p 1&u 2Fnetahtm ... more details
is implemented as solid phase synthesis using phosphoramidite method and phosphoramidite building ... synthesis see Phosphoramidite building blocks below . ref name sinha Many later improvements to the manufacturing of building blocks, instrumentation, and synthetic protocols made the phosphoramidite .... ref name BSL Synthesis by the phosphoramidite method Building blocks Nucleoside phosphoramidites main Nucleoside phosphoramidite File phosphoramidite1.png thumb 500px Protected 2 deoxynucleoside ... of internucleosidic linkages is dramatically improved by using 3 O N , N diisopropyl phosphoramidite ... r15 ref name biosearch ref name chemgenes ref name r17 and most common nucleoside phosphoramidite building ... reaction. However, a phosphoramidite derived from the N2 unprotected 5 O DMT 2 deoxyguanosine is poorly .... ref name sinha Once a phosphoramidite has been coupled to the solid support bound oligonucleotide ... are the phosphoramidite reagents designed to introduce various functionalities at the termini of synthetic ... groups, one of which is often protected with the DMT group while the other bears the reactive phosphoramidite .... A very short selection of commercial phosphoramidite reagents is shown in Scheme for the demonstration ... of oligonucleotides by phosphoramidite method. Oligonucleotide synthesis is carried out ... of the desired full length product. Step 2 Coupling A 0.02 0.2 M solution of nucleoside phosphoramidite ... as activator for 2 O TBDMS phosphoramidite building blocks in RNA synthesis volume 43 issue 5 pages ... containing solid support. The activated phosphoramidite in 1.5 20 fold excess over ... phosphoramidite moiety of the incoming nucleoside phosphoramidite to form a phosphite triester linkage. The phosphoramidite coupling is very rapid and requires, on small scale, about 20 ... and, in the phosphoramidite method, serves two purposes. After the completion of the coupling ... phase synthesis of oligonucleotides using phosphoramidite derivatives volume 14 issue 16 pages 6453 ... more details
Phosphoramidite building blocks oligonucleotide synthesis . Methoxymethyl ether MOM Removed by acid ... synthesis Phosphoramidite building blocks oligonucleotide synthesis for protection of N4 ... amides. Benzoyl Bz Anchor Bz group is common in Oligonucleotide synthesis Phosphoramidite building ... Phosphoramidite building blocks oligonucleotide synthesis . Methyl Me Anchor Me removed by strong ... more details
6 FAM phosphoramidite . Fluorescein also has an isosbestic point equal absorption for all pH pH values ... FITC FAM.png thumb left 400px Fluorescein isothiocyanate and 6 FAM phosphoramidite There are many ... , several phosphoramidite reagents containing protected fluorescein, e.g. fluorescein amidite 6 FAM phosphoramidite 2 , ref Brush, C. K. Fluorescein Labelled Phosphoramidites. US Patent 5,583,236. http ... more details
File Acrydite.svg thumb right Acrydite Acrydite is a phosphoramidite that allows the synthesis of oligonucleotides with a Methacrylic acid methacryl group at the 5 end less commonly 3 or internal . Acryl oligonucleotides have been tested, but the acryl group is not stable to storage. Acrydite modified oligonucletides can react with nucleophile s such as thiol s Michael addition chemistry , this forms the basis of the ez rays chemistry which was used for microarray s. More importantly, Acrydite modified oligonucleotides can be incorporated, stoichiometrically, into hydrogels such as polyacrylamide , using standard free radical polymerization chemistry, where the double bond in the Acrydite group reacts with other activated double bond containing compounds such as acrylamide . History The idea of acrylamide modified DNA was developed by T. Christian Boles, while working at Mosaic Technologies, a now defunct biotechnology company located in Waltham, MA. The IP was licensed, along with a microarray technology ez rays to Matrix Technologies, of Hudson, NH, which is now part of Thermo Scientific. Acrydite modified oligonucleotides can be obtained from vendors such as Integrated DNA Technologies IDT . Hybridgel The first use of Acrydite was in a technology called hybridgel. ref Biotechniques. 1998 Sep 25 3 516 21. Mutation typing using electrophoresis and gel immobilized Acrydite probes. Kenney M, Ray S, Boles TC. ref In Hybridgel, Acrydite modified oligos are incorporated into a standard polyacrylamide gel system as complementary ss nucleic acid moves past the immobilized Acrydite oligos, the complementary DNA is captured. Hybridgel like technology is widely used as a DNA purification system, as in a DNA sequencing technology developed at Berkeley. ref Proc Natl Acad Sci U S A. 2006 May 9 103 19 7240 7245. Microfabricated bioprocessor for integrated nanoliter scale Sanger DNA sequencing ref Acrydite technology can also be used to purify DNA, as in the Pre gen colon cancer ... more details
in the course of oligonucleotide synthesis by the phosphoramidite method using commercial biotin phosphoramidite. ref cite journal author Pon, R. T. journal Tetrahedron Lett. year 1991 volume 32 issue 14 pages 1715 1718 doi 10.1016 S0040 4039 00 74311 5 title A long chain biotin phosphoramidite ... more details
In chemistry a chiral ligand is a specially adapted ligand used for asymmetric synthesis . This ligand is an enantiopure organic compound which combines with a metal center by chelation to form an asymmetric catalyst . This catalyst engages in a chemical reaction and transfers its chirality to the reaction product which as a result also becomes chiral. In an ideal reaction one equivalent of catalyst can turnover number turn over many more equivalents of reactant which enables the synthesis of a large amount of a chiral compound from achiral precursors with the aid of a very small often expensive chiral ligand. First discovery The first such ligand, the phosphine diphosphine DiPAMP was developed in 1968 by William S. Knowles of Monsanto Company , who won the 2001 Nobel Prize in Chemistry , ref Nobel prize 2001 www.nobelprize.org http nobelprize.org nobel prizes chemistry laureates 2001 public.html Link ref and ultimately used in the industrial production of L DOPA . Image L DOPA synthesis2.png 550px Asymmetric L DOPA synthesis Privileged ligands Many thousands of chiral ligands have been prepared and tested since then but only several compound classes have been found to have a general scope. These ligands are therefore called privileged ligands . ref Design of chiral ligands for asymmetric catalysis From C2 symmetric P,P and N,N ligands to sterically and electronically nonsymmetrical P,N ligands Andreas Pfaltz and William J. Drury III PNAS, April 20, 2004 vol. 101 no. 16 5723 5726 DOI 10.1073 pnas.0307152101 ref ref Privileged Chiral Catalysts Tehshik P. Yoon, Eric N. Jacobsen Science 14 March 2003 Vol. 299. no. 5613, pp. 1691 1693 DOI 10.1126 science.1083622 PMID 12637734 ref Important members depicted below are BINOL , BINAP , TADDOL , DIOP , bisoxazoline ligand BOX and DuPhos a phosphine ligand , all available as enantiomeric pairs. Image ChiralLigands.png 400px A selection of chiral ligands Other members are Salen ligand Salen , cinchona alkaloid s and phosphoram ... more details
coord 42.346397 71.101853 display title The Molecular Biology Core Facilities MBCF was created to allow investigators at the Dana Farber Cancer Institute DFCI access to cutting edge molecular biology tools which would be tested and developed in a shared setting. Collaborations can be set up with anyone in the world. Although these services are primarily focused on Cancer and AIDS research, there is a broad spectrum of research that uses these resources. ref cite web url http cbmi.catalyst.harvard.edu cores cat core.html?core id 67&category id 8&navMode cat title The Molecular Biology Core Facilities at Dana Farber Cancer Institute Harvard Cores accessdate 18 April 2009 publisher Harvard Catalyst ref Image Mbcflogo.png thumb right The MBCF logo. History The http mbcf.dfci.harvard.edu MBCF at DFCI was first started in 1984 to supply small oligonucleotide s to researchers. Marvin Caruthers at the University of Colorado determined how to synthesize small oligonucleotides by solid phase synthesis using the phosphoramidite method. ref Beaucage SL, Caruthers MH. 1981 Tetrahedron Lett. 22, 1859 62. ref ref McBride LJ, Caruthers MH. 1983 Tetrahedron Lett. 24, 245 8. ref Carruthers and Leroy Hood at the California Institute of Technology constructed the first automated Solid phase synthesis DNA Synthesizer which the MBCF successfully tested after discussions with Don Taylor. Because of the growing demand for oligonucleotide primers to initiate DNA replication and for probes, a plan was put into place to develop a core facility to produce reagents for molecular biologists as well as instrumentation for the analysis of DNA and protein samples. This plan stated that a charge back method would be put in place to fairly spread the resources as a shared facility. A Solid phase synthesis Peptide Synthesizer using Bruce Merrifield s solid phase peptide synthesis ref cite journal author R. B. Merrifield title Solid Phase Peptide Synthesis. I. The Synthesis of a Tetrapeptide year 1963 ... more details
Distinguish degenerate bases For phosphoramidite synthesis of nucleic acids Oligonucleotide synthesis File Difference DNA RNA EN.svg thumb 300px RNA with its nucleobase s to the left and DNA to the right. Nucleic acid analogues are compounds structurally similar Analog chemistry analog to naturally occurring RNA and DNA , used in medicine and in molecular biology research. Nucleic acid s are chains of nucleotides, which are composed of three parts a phosphate backbone, a pucker shaped pentose sugar, either ribose or deoxyribose , and one of four nucleobase s. An analogue may have any of these altered. Typically the analogue nucleobases confer, among other things, different base pairing and base stacking proprieties. Examples include universal bases, which can pair with all four canon bases, and phosphate sugar backbone analogues such as peptide nucleic acid PNA , which affect the properties of the chain PNA can even form a triple helix . ref Petersson B et al. Crystal structure of a partly self complementary peptide nucleic acid PNA oligomer showing a duplex triplex network. J Am Chem Soc. 2005 Feb 9 127 5 1424 30. ref Artificial nucleic acids include peptide nucleic acid PNA , Morpholino and locked nucleic acid LNA , as well as GNA nucleic acid glycol nucleic acid GNA and TNA nucleic acid threose nucleic acid TNA . Each of these is distinguished from naturally occurring DNA or RNA by changes to the backbone of the molecule. Medicine main Nucleoside analogues Several nucleoside analogues are used as antiviral or anticancer agents. The Virus viral polymerase incorporates these compounds with non canon bases. These compounds are activated in the cells by being converted into nucleotides, they are administered as nucleosides since charged nucleotides cannot easily cross cell ... using Oligonucleotide synthesis phosphoramidite strategy or, for PNA, methods of peptide ... of S br 2 amino 6 2 thienyl purine S br File Phosphoramidite.png 200px phosphoramidite br Phosphoramidite ... more details
using nucleotides, called phosphoramidite s, normal nucleotides which have protection groups preventing amine, hydroxyl groups and phosphate groups interacting incorrectly. One phosphoramidite ... more details
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