Method of detecting micrometastasis

Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid

Reexamination Certificate

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C536S024300, C435S091200

Reexamination Certificate

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10839202

ABSTRACT:
A method of detecting micrometastasis of tumor cells in a sample obtained from a region of a subject other than the primary focus of the tumor by using a first primer complementary to part of a specific sequence in the RNA from tumor cells and a second primer homologous to part of the specific sequence (either of which additionally has a promoter sequence for an RNA polymerase at the 5′ end), which comprises (1) synthesizing a cDNA by the action of an enzyme having RNA-dependent DNA polymerase activity by using the specific sequence as a template, (2) degrading the RNA strand in the RNA-DNA double strand by an enzyme having a ribonuclease H activity (to give a single-stranded DNA), (3) forming a double-stranded DNA having a promoter sequence which can be transcribed into an RNA homologous or complementary to the specific sequence by using the single-stranded DNA as a template by the action of an enzyme having DNA-dependent DNA polymerase activity, and (4) transcribing the double-stranded DNA into an RNA transcript (which acts as a template in the subsequent cDNA synthesis in the reaction (1)) by the action of an enzyme having RNA polymerase activity and detecting the mRNA.

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