fC-CET
5fC Chemical Labeling and C-to-T Transition During PCR
fC-CET is a bisulfite-free method for whole-genome analysis of 5fC. The method is based on selective chemical labeling of 5fC and subsequent C-to-T transition during PCR (Xia et al., 2015). In this method, gDNA is labeled sequentially with an azido derivative of 1,3-indandione (AI) and then conjugated to biotin using click chemistry. The gDNA is enriched by streptavidin pull-down and ligated to adapters, followed by PCR and sequencing. C-to-T transitions during PCR amplification are searched specifically to define 5fC sites in the genome.
Advantages:
- Results are in agreement with fC-Seal
- Sequences with a single 5fC are enriched ~100-fold
- No noticeable DNA degradation; ideal for analyses of precious DNA samples
Disadvantages:
- Provides only relative enrichment of cytosines marked by 5fC (Wu et al., 2016)
Reagents:
Illumina Library prep and Array Kit Selector
Reviews:
None available yet
References:
Xia B., Han D., Lu X., et al. Bisulfite-free, base-resolution analysis of 5-formylcytosine at the genome scale. Nat Methods. 2015;12:1047-1050