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A single cell was lysed with 100 ng of the carrier RNA mixture. The sequencing library was constructed following the manufacturer’s protocol using a NEBNext kit (E7420). After adaptor ligation, the cDNA fragments from the RNA carrier were removed by Not I digestion. After Not I digestion, the library DNA was purified for USER digestion and PCR amplification. PCR amplification was performed using primers containing 8-nt degenerate barcodes to perform the first 10 cycles of library amplification, followed by another 15 cycles of amplification by regular P5/P7 primers. The directional Holo-Seq library of total RNA was used for target capturing using the Illumina TruSeq Exome Capture kit (FC-144-1004). |
Targeted-Capture |
TRANSCRIPTOMIC |
padlock probes capture method |
SINGLE
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