Device and process for assays using binding members
A technology that combines components and covers components, applied in the field of analysis, and can solve problems such as difficulties
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Embodiment 1
[0425] Example 1: Competitive Assay for Determination of Human Poliovirus 1 DNA
[0426] The schematic diagram of the performed competitive analysis is shown in Figure 22 middle. The DNA of human poliovirus 1 isolate TCDC01-861 (GenBank accession number AF538843) was cloned into an appropriate expression vector (pCR 2.1-TOPO , Clontech, Inc. Palo Alto, CA, USA), used as a DNA template (also referred to herein as "EV" (enterovirus) DNA).
[0427] contains 10 for each of the two 4 Samples of three DNA copies were analyzed in parallel: the first sample was PCR amplified using a Rotor-Gene 6000 real-time rotary PCR analyzer (Corbett Life Sciences, Sydney, Australia) according to the manufacturer's instructions.
[0428] The second sample also contains a Taqman The probe has the same nucleotide sequence, but contains a CY3 carbocyanine label (Invitrogen Corporation, Carlsbad, CA, USA) at its 3' end instead of a FAM / TAMRA-labeled reporter molecule, and is analyzed in a re...
Embodiment 2
[0445] Example 2: Array-Based Competitive Assay for Determination of HIV1 gag / env DNA
[0446] The schematic diagram of the competitive analysis performed is shown in Figure 24 In A. The DNA of the synthetic HIV1 gag / env fusion construct (EMBL Accession No. A06258) was cloned into the expression vector pCR 2.1-TOPO (Clontech, Inc. Palo Alto, CA, USA) EcoRI endonuclease restriction sites were used as DNA templates.
[0447] Additionally, the following PCR primers were used:
[0448] Forward PCR primers:
[0449] cdia: 5′-TGAAGGGTACTAGTAGTTCCTGCTATGTC-3′
[0450] (corresponding to nucleotides 214-232 of A06258)
[0451] Reverse PCR primers:
[0452] cdis: 5′-ATCAAGCAGCCATGCAAATGTT-3′
[0453] (corresponding to nucleotides 384-405 of A06258)
[0454] Thus, PCR resulted in the amplification of a 151 bp DNA fragment having the following sequence: 5'-ATC AAG CAG CCA TGC AAA TGT TAA AAG AGA CCA TCAATG AGG AAG CTG CAG AAT GGG ATA GAT TGC ATC CAG TCCATG GAG GGC CTA TTG C...
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