ddNTPDideoxynucleotide Triphosphate
ddNTPDideoxyribonucleotide triphosphate
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For the amplification of the targeted PCA3 and PSA amplicons and the measurements of PCA3/PSA ratio the following PCR conditions were applied: master mix with final volume 10 [micro]l: 2 [micro]l cDNA input; 0.2 [micro]l of specific PCA3 and PSA primers with a concentration 20 pmol/[micro]l; 1 [micro]l ddNTP (5 mM) and 1 [micro]l 10X Buffer; 0.05 [micro]l Taq DNA Polymerase 250 U 5U/[micro]l (GeNet Bio, Korea) and 5.15 [micro]l PCR water.
In a Sanger sequencing reaction, then, we end up with a mixture of polymerase extension products in which a very small fraction of the new DNA strands--all derived from the same template and starting location as defined by the primer--have selectively terminated at each base position along the template; and the terminating ddNTP identifies its template partner by the rules of hybridization mentioned above.
The oligonucleotides (Table 2) related to K1637K and S1647T were synthesized to verify the feasibility of the ddNTP blocking efficiency.
(36) Dideoxynucleotide sequencing is based on the synthesis of new DNA strands (complementary to the target sequence of interest) in the presence of primers, deoxynucleotide triphosphates (dNTPs) and dideoxynucleotide triphosphate chain terminators (ddNTPs).
[4] Nonstandard abbreviations: SNP, single-nucleotide polymorphism; SPC, solid-phase capture; SBE, single-base extension; CYP450, cytochrome P450; NCBI, National Center for Biotechnology Information; ddNTP, dideoxyribonucleotide triphosphate.
Primer extension was performed using a SNaPshot ddNTP Primer Extension Kit (Applied Biosystems, Foster City, CA).
(1) Nonstandard abbreviations: HAEC, human aortic endothelial cell; HUVEC, human umbIIical vein endothelial cell; dNTP, deoxynucleoside triphosphate; ddNTP, dideoxynucleoside triphosphate; Ct, threshold cycle; s/n, signal-to-noise rafio; and s/i, signal iracfion.
Primer extension reactions were performed using the SNaPshot ddNTP Primer Extension Kit (Applied Biosystems, Foster City, CA, USA).
Primer extension reactions were conducted using the SNaPshot ddNTP Primer Extension Kit (Applied Biosystems, Foster City, CA, USA).