The first nucleotide dispensed, dATP, is not incorporated into either nascent strand, whereas the next one, dGTP
, is incorporated twice on the strand for the wild-type allele and only once on the strand for the mutant allele, causing the polymerase molecules to go out of phase.
ABBREVIATIONS: bDNA = Branched DNA; DNA = Deoxyribonucleic Acid; dNTP = Deoxyribonucleotides; dATP = Deoxyadenosine Triphosphate; dCTP = Deoxycytidine Triphosphate; dGTP
= Deoxyguanosine Triphosphate; dTTP = Deoxythymidine Triphosphate; FDA = Food and Drug Administration; FRET = Fluorescence Resonance Energy Transfer; MRSA = Methicillin Resistant Staph aureus; MTB = Mycobacterium tuberculosis; NASBA = Nucleic Acid Sequence Based Amplification; PCR = Polymerase Chain Reaction; PFGE = Pulsed-Field Gel Electrophoresis; RT-PCR = Reverse Transcription Polymerase Chain Reaction; Taq = Thermus aquaticus; TMA = Transcription-Mediated Amplification; VRE = Vancomycin Resistant Enterococcus.
Omission of dATP, dTTP, or dGTP
during the telomerase extension step of the TRAP reaction resulted in no elongation products being amplified; however, omission of dCTP did not affect the formation of a ladder pattern of telomerase products (data not shown).
2 mM dATP, dCTP, dGTP
and dTTP), 1 U Taq DNA polymerase enzyme (Roche), 10 |iM of forward and reserve primers (Sigma-Genosys), and 10 ng of DNA template.
1 mM each of dATP, dGTP
, dTTP and dCTP, one unit of Taq DNA polymerase (Perkin Elmer, Norwalk, Conn.
2], 200 [micron]M de cada dGTP
, dATP, dTTP, dCTP (Promega, Madison, WI), O,5 [micron]M de cada iniciador (5'-ATCCAAGACAACACTA CTAA-3' y 5'-AAATATCCTCA AAGTTCC-3') y 2,5 unidades de Taq polimerasa (Corpogen, Colombia).
75 mM MgCl2, 350 mM each of dATP, dGTP
, dTTP, dCTP, 400 picomoles of primers (wsp-F and wsp-R), 1 unit of Pwo and 5 units of Taq DNA polymerases (Jeyaprakash & Hoy 2000).
1% Triton X-100], 1 unit of Taq polymerase (Banglore Genei), 200 uM (each) deoxynucleoside triphosphate (dATP, dCTP, dGTP
, and dTTP) (Banglore Genei), 10 pmol of each primer, 5 [mu]l of sample DNA, and ultra pure sterile water added to 25 [mu]l.
2], 200 [micro]M each dATP, dCTP, dGTP
and dTTP, 1 [micro]M each primer and 1 unit of DNA polymerase (BioTools Labs, Madrid, Spain) in a final volume of 25 [micro]l.
0 [micro]l dNTP [Roche] (10 mM of each deoxyribonucleotide: dATP, dCTP, dGTP
, and dTTP), 5.
2], dNTPs (dATP, dGTP
, dCTP, dTTP, 200 [micro]M each), and 5% DMSO.
2 mM each dATP, dCTP, dGTP
, and dTTP, 1 [micro]M each forward and reverse primer, 0.