التفاصيل البيبلوغرافية
| العنوان: |
A droplet microfluidic platform for efficient enzymatic chromatin digestion enables robust determination of nucleosome positioning. |
| المؤلفون: |
Xu, Yi1,2, Bailey, Ryan C.1,2,3, Lee, Jeong-Heon4, Ordog, Tamas3,4, Li, Zhaoyu4, Wang, Liguo4 |
| المصدر: |
Lab on a Chip. 9/7/2018, Vol. 18 Issue 17, p2583-2592. 10p. |
| مصطلحات موضوعية: |
*MICROFLUIDICS, *CHROMATIN, *DIGESTION, *EPIGENETICS, *ENZYMATIC analysis |
| مستخلص: |
The first step in chromatin-based epigenetic assays involves the fragmentation of chromatin to facilitate precise genomic localization of the associated DNA. Here, we report the development of a droplet microfluidic device that can rapidly and efficiently digest chromatin into single nucleosomes starting from whole-cell input material offering simplified and automated processing compared to conventional manual preparation. We demonstrate the digestion of chromatin from 2500–125 000 Jurkat cells using micrococcal nuclease for enzymatic processing. We show that the yield of mononucleosomal DNA can be optimized by controlling enzyme concentration and incubation time, with resulting mononucleosome yields exceeding 80%. Bioinformatic analysis of sequenced mononucleosomal DNA (MNase-seq) indicated a high degree of reproducibility and concordance (97–99%) compared with conventionally processed preparations. Our results demonstrate the feasibility of robust and automated nucleosome preparation using a droplet microfluidic platform for nucleosome positioning and downstream epigenomic assays. [ABSTRACT FROM AUTHOR] |
| قاعدة البيانات: |
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