The present application belongs to the field of biological sequencing technology, and discloses a MIP probe and its application in
urine free DNA capture and
library construction. The MIP probe includes, from 5'-3', a 5' capture arm, an F-
terminal adapter containing 10nt
hypoxanthine, an R-
terminal adapter, and a 3' capture arm. The
present method prepares
a DNA library by complementary base
pairing, extension, and connection of the capture arm, is compatible with
DNA fragments of different lengths and single-stranded
DNA, improves template utilization, and thus facilitates the detection of low-frequency mutations in
urine free DNA. The
present method fixes the length of the
library insert fragment by controlling the span length of the
chromosomal DNA to be tested between the capture arms on both sides of the probe, making the DNBSEQ platform sequencing
fluorescence signal more uniform, and the data output and quality higher. The
present method combines the three steps of capture, library construction, and
machine library preparation, eliminating the operations such as intermediate library
quality control and pre-capture
drying and concentration in conventional methods, and the
operation time is shorter; there is no PCR amplification during the capture and library construction process of the present method, which reduces the error accumulation due to PCR amplification, reduces
background noise, and improves the sensitivity and specificity of low-frequency
mutation detection.