Melting temperature of molecular beacons as an indicator of the ligase detection reaction for multiplex detection of point mutations

Abstract

A novel strategy based on the ligase detection reaction (LDR) using the melting temperature of molecular beacons as the indicator is presented for the multiplex detection of gene mutations.

Real-time detection of SARS-CoV-2 in clinical samples via ultrafast ligation-dependent RNA transcription amplification

“…Due to the high specicity, ligation-dependent methods have been used for the detection of pathogens and markers of genetic disorders. [18][19][20][21] However, both RNA transcription and the binding of uorogenic dye to the RNA aptamer take several hours, making the present ligationdependent methods time-consuming and undesirable for onsite rapid detection of emerging infectious diseases like COVID-19. Furthermore, most of the reported ligationdependent methods are based on end-point detection, which is generally considered to be of lower sensitivity and reliability than real-time detection.…”

Thermus thermophilus DNA Ligase Connects Two Fragments Having Exceptionally Short Complementary Termini at High Temperatures

“…T hermus thermophilus DNA ligase (Tth DNA ligase) catalyzes NAD-dependent phosphodiester bond formation, and has been utilized for ligase chain reaction (LCR), 1,2 ligase detection reaction (LDR), 3,4 and specific circularization for rolling circle amplification (RCA). 5−7 One of the most important characteristics of this ligase is the remarkable activity at high temperatures (45−75 °C), as compared with most other ligases active below 37 °C (e.g., T4 DNA ligase).…”

“…T hermus thermophilus DNA ligase ( Tth DNA ligase) catalyzes NAD-dependent phosphodiester bond formation, and has been utilized for ligase chain reaction (LCR), , ligase detection reaction (LDR), , and specific circularization for rolling circle amplification (RCA). − One of the most important characteristics of this ligase is the remarkable activity at high temperatures (45–75 °C), as compared with most other ligases active below 37 °C (e.g., T4 DNA ligase). − High ligation temperature minimizes nonspecific ligation, and facilitates unique applications which cannot be otherwise accomplished. On the other hand, for ligation by the Tth DNA ligase, long complementary single-stranded DNA (ssDNA) parts (e.g., >30 bp) were usually required to place the two termini for ligation in close proximity due to the difficult hybridization at high temperatures.…”