| 建库信息 |
| 文库名称 |
文库构建方法 |
建库策略 |
样品来源 |
实验选择 |
文库布局 |
|
Tissue was crosslinked by 40 ml 2% formaldehyde solution for 15 minutes and subsequently added Glycine to quench the crosslinking reaction. Crosslinked tissue was ground up with liquid nitrogen and nuclei were extracted by centrifuge for three times which included filtration at 4000 rpm and 4Celsius for 20 min and extraction at 14,000 rpm and 4Celsius for 10 min. The chromatin was then solubilized with dilute SDS and incubated for 10 minutes at 65Celsius. After quenching the SDS with Triton X-100, restriction enzyme (400 units MboI) was administered overnight at 37Celsius on a rocking platform. The ends of the DNA were labeled by biotin-14-dCTP (Invitrogen, 19518-018) and were linked for crosslinked fragments. After proximal chromatin DNA was religated, the chromatins were reversed crosslinked by incubating with proteinase K (Invitrogen, 25530-031) at 65Celsius and then extracted DNA using phenol-chloroform. T4 DNA polymerase (NEB, M0203L) was used to discard biotin-C from non-ligated fragment ends. Sonication was used to cleave fragments to a size of 100-500 base pairs. T4 DNA polymerase, T4 polynucleotide kinase (NEB, M0201), and Klenow DNA polymerase were used to repair the fragment ends. Streptavidin magnetic beads were used to enrich biotin-tagged target fragments. The fragment ends were A-tailed by Klenow (NEB, M0212L) and then ligated using an Illumina paired-end sequencing adaptor (Illumina, PN 1001782, 1001783). Hi-C libraries were constructed following Illumina protocols and sequenced using PE150 reads with a sequenced in Illumina HiSeq instruments. |
Hi-C |
GENOMIC |
RANDOM |
PAIRED
|
|
