HpaII

HpaII is a type II restriction endonuclease that recognizes the DNA sequence 5'-CCGG-3' and cleaves within or near this site to generate cohesive ends. It is derived from Haemophilus species and is commonly used in molecular biology for DNA digestion and epigenetic studies due to its sensitivity to cytosine methylation at the recognition sequence.

HpaII is methylation-sensitive. It is inhibited by methylation of the inner cytosine at the CCGG site, such that methylation of the second cytosine prevents cutting. By contrast, its isoschizomer MspI recognizes the same sequence but is blocked by methylation of the outer cytosine. This differential sensitivity allows researchers to infer methylation patterns by comparing digestion with HpaII and MspI, a principle exploited in methylation-sensitive assays.

Applications of HpaII include restriction fragment analysis, development of methylation-sensitive molecular markers, and epigenetic studies. It is frequently used in conjunction with MspI in techniques such as MS-AFLP (methylation-sensitive amplified fragment length polymorphism) to assess CpG methylation status across genomes. While newer methods such as bisulfite sequencing can provide higher-resolution data, HpaII remains a valuable, cost-effective tool for examining methylation-dependent DNA restriction.

In summary, HpaII is a methylation-sensitive restriction enzyme that recognizes CCGG and helps distinguish methylation states at CpG dinucleotides, aiding both basic molecular biology workflows and epigenetic analyses.