Methods: We tested a panel of multiplexed, high-throughput sequenced introns in the small mammal communities of two UNESCO World Heritage Sites on different continents to assess their viability for large-scale monitoring of genetic variability in a spectrum of diverse species. To enhance applicability across other systems, the bioinformatic pipeline for primer design was outlined.
Results: The number of loci amplified and amplification evenness decreased as phylogenetic distance increased from the reference taxa, yet several loci were still variable across multiple mammal orders.
Conclusions: Genetic variability found is informative for population genetic analyses and for addressing phylogeographic and phylogenetic questions, illustrated by small mammal examples here.
Materials and methods: We analysed 100 unrelated HA and 15 unrelated HB patients for genetic alterations in the F8 and F9 genes by using the long-range PCR, DNA sequencing, and the multiplex-ligation-dependent probe amplification assays. The prediction software was used to confirm the effects of these mutations on factor VIII and IX proteins.
Results: 44 (53%) of the severe HA patients were positive for F8 intron 22 inversion, and three (3.6%) were positive for intron one inversion. There were 22 novel mutations in F8, including missense (8), frameshift (9), splice site (3), large deletion (1) and nonsense (1) mutations. In HB patients, four novel mutations were identified including the splice site (1), small deletion (1), large deletion (1) and missense (1) mutation.
Discussion: The mutational spectrum of F8 in Malaysian patients is heterogeneous, with a slightly higher frequency of intron 22 inversion in these severe HA patients when compared to other Asian populations. Identification of these mutational profiles in F8 and F9 genes among Malaysian patients will provide a useful reference for the early detection and diagnosis of HA and HB in the Malaysian population.