Herculase 2 fusion enzyme kit

Manufactured by Agilent Technologies

The Herculase II Fusion Enzyme Kit is a high-fidelity DNA polymerase designed for PCR amplification. It provides reliable and accurate DNA amplification for a variety of applications.

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Lab products found in correlation

Ni nta agarose, by Thermo Fisher Scientific (1 mentions) Flag m2 monoclonal antibody, by Merck Group (1 mentions) Mg132, by Merck Group (1 mentions) Cycloheximide (chx), by Merck Group (1 mentions) Blocking agent, by Agilent Technologies (1 mentions) Kapa library quantification kit, by Illumina (1 mentions) Hybridization buffer, by Agilent Technologies (1 mentions) Human cot 1, by Thermo Fisher Scientific (1 mentions) Salmon sperm dna, by Thermo Fisher Scientific (1 mentions)

2 protocols using herculase 2 fusion enzyme kit

Functional Study of CRY2 T300A Mutant

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PCR-amplified human CRY2 was cloned into PCMV5 or PDEST15. CRY2 T300A mutant was made by the Herculase II Fusion Enzyme Kit (Agilent Technologies). siRNA oligos were designed and manufactured by Ruibo (Guangzhou, China), targeting CRY2, sense: 5′-GAACGAAUGAAGCAGAUUU. Flag-FBXW7 was previously described (31 (link)). Cycloheximide and MG132 were obtained from Sigma. Ni-NTA Agarose was obtained from Invitrogen (#R901-15). Antibodies: Flag (M2 monoclonal antibody, Sigma, F3165), CRY2 (Abcam), myc (Santa cruz, sc-40).

Fang L., Yang Z., Zhou J., Tung J.Y., Hsiao C.D., Wang L., Deng Y., Wang P., Wang J, & Lee M.H. (2015). Circadian clock gene CRY2 degradation is involved in chemoresistance of colorectal cancer. Molecular cancer therapeutics, 14(6), 1476-1487.

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Target Capture Probe Enrichment Protocol

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Target capture probes were used to enrich libraries using in-solution Hybridization [16 ,28 ]. Each barcoded library was mixed with 2x hybridization buffer (Agilent Technologies), 10x blocking agent (Agilent Technologies), blocking oligonucleotides, human Cot-1 (Invitrogen, Carlsbad, CA, USA) and salmon sperm DNA (Invitrogen) [16 ,25 (link),28 ,29 ]. The libraries were then incubated with the biotinylated capture probes for 48 hours at 66°C and were eluted by heating [16 ,27 ]. Enriched fetal-specific regions were amplified for 12 cycles using Herculase II Fusion Enzyme kit (Agilent Technologies) and outward bound adaptor primers [27 ]. Following quantification with the KAPA Library Quantification Kit Illumina (KAPA Biosystems, Boston, MA, USA) the amplified post-captured libraries were sequenced on a HiSeq 2500 sequencing system platform (Illumina, San Diego, USA).

Keravnou A., Ioannides M., Loizides C., Tsangaras K., Achilleos A., Mina P., Kypri E., Hadjidaniel M.D., Neofytou M., Kyriacou S., Sismani C., Koumbaris G, & Patsalis P.C. (2018). MeDIP combined with in-solution targeted enrichment followed by NGS: Inter-individual methylation variability of fetal-specific biomarkers and their implementation in a proof of concept study for NIPT. PLoS ONE, 13(6), e0199010.

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