TY - JOUR
T1 - Rapid and unbiased extraction of chromatin associated RNAs from purified native chromatin
AU - Zhou, Zhongwu
AU - Yang, Yi
AU - Konieczny, Stephen F.
AU - Irudayaraj, Joseph M.K.
N1 - This project was partly funded by the W.M. Keck Foundation . USDA-ARS project number 935-42000-049-00D in conjunction with the Center for Food Safety Engineering at Purdue University and the Indiana Clinical and Translational Sciences Institute funded, in part by Grant Number ( TR000006 ) from the NIH are acknowledged.
This project was partly funded by the W.M. Keck Foundation. USDA-ARS project number 935-42000-049-00D in conjunction with the Center for Food Safety Engineering at Purdue University and the Indiana Clinical and Translational Sciences Institute funded, in part by Grant Number (TR000006) from the NIH are acknowledged.
PY - 2015/12/24
Y1 - 2015/12/24
N2 - An ultra fast and unbiased method that uses salicylic acid coated magnetic nanoparticles (SAMNPs) and magnetophoretic chromatography is developed to extract chromatin associated RNAs (CARs). The SAMNPs were first used for enriching cells from the cell culture media and further used for capturing chromatin after cells were lysed. The formed SAMNPs-chromatin complexes were transferred to a viscous polyethylene glycol (PEG) solution stored in a 200-μl pipette tip. Due to the difference in viscosities, a bi-layer liquid was formed inside the pipette tip. The SAMNPs-chromatin complexes were separated from the free SAMNPs and free RNA-SAMNPs complexes by applying an external magnetic field. The CARs were further extracted from the SAMNP-chromatin complexes directly. The extracted CARs were reverse transcribed as cDNA and further characterized by real-time qPCR. The total assay time taken for cell separation, chromatin purification and chromatin associated RNAs extraction can be accomplished in less than 2 h.
AB - An ultra fast and unbiased method that uses salicylic acid coated magnetic nanoparticles (SAMNPs) and magnetophoretic chromatography is developed to extract chromatin associated RNAs (CARs). The SAMNPs were first used for enriching cells from the cell culture media and further used for capturing chromatin after cells were lysed. The formed SAMNPs-chromatin complexes were transferred to a viscous polyethylene glycol (PEG) solution stored in a 200-μl pipette tip. Due to the difference in viscosities, a bi-layer liquid was formed inside the pipette tip. The SAMNPs-chromatin complexes were separated from the free SAMNPs and free RNA-SAMNPs complexes by applying an external magnetic field. The CARs were further extracted from the SAMNP-chromatin complexes directly. The extracted CARs were reverse transcribed as cDNA and further characterized by real-time qPCR. The total assay time taken for cell separation, chromatin purification and chromatin associated RNAs extraction can be accomplished in less than 2 h.
KW - Heterochromatin
KW - Magnetic nanoparticle
KW - Magnetophoretic chromatography
KW - Soluble chromatin
KW - Unbiased preparation of chromatin associated RNAs
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U2 - 10.1016/j.chroma.2015.11.067
DO - 10.1016/j.chroma.2015.11.067
M3 - Article
C2 - 26643718
AN - SCOPUS:84983184611
SN - 0021-9673
VL - 1426
SP - 64
EP - 68
JO - Journal of Chromatography A
JF - Journal of Chromatography A
ER -