Hi-C of Babesia microti: Bm1438 synchronized tetrads replicate 2
Cell cultures were crosslinked for 25 minutes at 37C in 1.25% formaldehyde in a total volume between 1 and 10 ml, depending on the number of parasites harvested. The reaction was quenched for 15 min at 37C, followed by extensive washes to remove cellular debris. To map the inter- and intrachromosomal contact counts, crosslinked parasites were subjected to the in situ Hi-C prodecure, using MboI for restriction digests. Libraries were generated using NEBNext Multiplex Oligos for Illumina and Kapa HiFi Hotstart ReadyMix (Kapa Biosystems) and amplified for a total of 12 PCR cycles (15s at 98C, 30s at 55C, 30s at 62C) and were subsequently sequenced using the Illumina HiSeq2500 platform, generating 50 bp paired-end reads.