Optimization of long-range PCR protocol to prepare filaggrin exon 3 libraries for PacBio long-read sequencing.

ABSTRACT:

Background

The filaggrin (FLG) protein, encoded by the FLG gene, is an intermediate filament-associated protein that plays a crucial role in the terminal stages of human epidermal differentiation. Loss-of-function mutations in the FLG exon 3 have been associated with skin diseases. The identification of causative mutations is challenging, due to the high sequence homology within its exon 3 (12,753 bp), which includes 10 to 12 filaggrin tandem repeats. With this study we aimed to obtain the whole FLG exon 3 sequence through PacBio technology, once 13-kb amplicons have been generated.

Methods and results

For the preparation of SMRTbell libraries to be sequenced using PacBio technology, we focused on optimizing a 2-step long-range PCR protocol to generate 13-kb amplicons coveri

SUBMITTER: Mareso C 

PROVIDER: S-EPMC10042914 | biostudies-literature | 2023 Apr

REPOSITORIES: biostudies-literature