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Abstract

Nucleotide sequence information derived from DNA segments of the human and other genomes is accumulating rapidly. However, it frequently proves difficult to use such short DNA segments to identify clones in genomic libraries or fragments in blots of the whole genome or for in situ analysis of chromosomes. Oligonucleotide probes, consisting of two target-complementary segments, connected by a linker sequence, were designed. Upon recognition of the specific nucleic acid molecule the ends of the probes were joined through the action of a ligase, creating circular DNA molecules catenated to the target sequence. These probes thus provide highly specific detection with minimal background.
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... Both MolBoolean and in situ PLA detected a To validate that the dual-colored RCA products observed in Ecadherin-β-catenin stains contain both tags and are not just individual single-colored RCPs located in close proximity, we designed padlock probes targeting the MolBoolean proximity probes (see Methods for design). Padlock probes consist of two target-complementary segments, connected by a linker sequence 18 . Upon recognition of the target DNA sequence (Fig. 4a), the 5′ and the 3′ end of the padlock probe can be joined by ligation with T4 ligase (Fig. 4b), creating a circular DNA molecule that is amplifiable by RCA 18 (Fig. 4c). ...
... Padlock probes consist of two target-complementary segments, connected by a linker sequence 18 . Upon recognition of the target DNA sequence (Fig. 4a), the 5′ and the 3′ end of the padlock probe can be joined by ligation with T4 ligase (Fig. 4b), creating a circular DNA molecule that is amplifiable by RCA 18 (Fig. 4c). As a result, regardless of the target proteins' proximity, each padlock probe always generates its own individual fluorescent signal and never a dual signal (Fig. 4d). ...
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