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The DNA degradation and contamination of the extracted DNA was monitored on 1% agarose gels. DNA purity was then detected using NanoDrop™ One UV-Vis spectrophotometer (Thermo Fisher Scientific, USA), of which OD260/280 ranging from 1.8 to 2.0 and OD 260/230 is between 2.0-2.2. At last, DNA concentration was further measured by Qubit® 4.0 Fluorometer (Invitrogen, USA).SMRTbell target size libraries were constructed for sequencing according to PacBio’s standard protocol (Pacific Biosciences, CA, USA) using either 10 kb or 20 kb preparation solutions. The main steps for library preparation are: (1) gDNA shearing, (2) DNA damage repair, (3) blunt-end ligation with hairpin adapters from the SMRTbell Template Prep Kit 1.0 (Pacific Biosciences), (4) size selection, and (5) binding to polymerase. Briefly, a total amount of 5 µg DNA per sample was used for the DNA library preparations. |
RNA-Seq |
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RT-PCR |
SINGLE
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