Fluorescent DNA Synthesis and Purification

The fluorescent DNA containing 20 Ronin-binding motifs was synthetized by IDT and cloned into the pUC19 vector using HiFi Assembly (NEB).^{39 (link)} Cy5-labeled M13(21) (/5Cy5/TGTAAAACGACGGCCAGT) and M13 reverse (/5Cy5/CAGGAAACAGCTATGAC) primers were used to amplify the synthetic DNA sequence by PCR, yielding a fluorescently labeled PCR product. The fluorescent PCR products were purified twice, first with the QIAGEN PCR purification kit and then with the NEB Monarch PCR purification kit.

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Dejosez M., Dall’Agnese A., Ramamoorthy M., Platt J., Yin X., Hogan M., Brosh R., Weintraub A.S., Hnisz D., Abraham B.J., Young R.A, & Zwaka T.P. (2023). Regulatory architecture of housekeeping genes is driven by promoter assemblies. Cell reports, 42(5), 112505.

Publication 2023

HiFi Assembly Monarch PCR purification kit

Dna sequence Primers Vector

Corresponding Organization : Whitehead Institute for Biomedical Research

Other organizations : St. Jude Children's Research Hospital

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Variable analysis

independent variables

Ronin-binding motifs in the fluorescent DNA

dependent variables

Amplification of the synthetic DNA sequence by PCR
Purification of the fluorescent PCR products

control variables

PUC19 vector used for cloning the fluorescent DNA
Cy5-labeled M13(21) and M13 reverse primers used for PCR amplification
QIAGEN PCR purification kit and NEB Monarch PCR purification kit used for purification

controls

No positive or negative controls were explicitly mentioned in the provided information.

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