The First Molecular Profiling of JAK2 Exon 12 and Flanking Intronic Sequences in Indonesia

Data on JAK2 Exon 12 mutations in Indonesia are not available. This study aimed to explore the presence of known JAK2 exon 12 coding mutations in V617F-negative myeloproliferative neoplasm (MPN) patients in Indonesia and to describe any novel intronic changes downstream of exon 12.

A total of 198 V617F-negative MPN samples were screened using allele-specific polymerase chain reaction (AS-PCR) for four commonly reported canonical exon 12 mutations. Of these, 57 cases were selected at random or for atypical PCR band patterns and underwent bidirectional Sanger sequencing of a 496-bp fragment covering exon 12 plus ~250 bp of the adjacent region. Variants were cross-referenced with gnomAD v4.1.0. Furthermore, SpliceAI was used to predict splice site gain or loss.

No coding mutations were identified in exon 12. However, two intronic variants were detected adjacently downstream: NC_000009.12:g.5070231_5070235delTCTTA in 29 of 57 cases (50.9%), which was not present in the gnomAD database, and NC_000009.12:g.5070319C>T in 1 of 57 cases (1.8%), corresponding to a rare gnomAD entry. The SpliceAI Δ scores remained below 0.2, a high-sensitivity threshold, indicating no predicted impact on canonical splicing.

Classic exon 12 coding mutations are apparently absent in Indonesian V617F-negative MPNs. We report a highly frequent downstream intronic deletion not recorded in global databases, as well as a rare single nucleotide variant (SNV) that aligns with gnomAD data. Although in silico tools suggest minimal splice disruption, RNA-level assays are needed to rule out the subtle effects of the variant. Our workflow, which combines allele-specific PCR and targeted sequencing, may be suitable for laboratories facing resource constraints.