Nd 1000 spectrophotometer

Sequencing analysis of ELOVL4 and PRPH2 was performed using the MiSeq NGS platform (Illumina, Inc., San Diego, CA, USA). The concentration and purity of DNA samples was quantified using an ND-1000 spectrophotometer (NanoDrop, Wilmington, DE, USA). A concentration of 50 ng/µL was used for the analysis. Exons 1-6 and 1-3 of ELOVL4 and PRPH2, respectively, and their flanking splice site junctions were amplified using polymerase chain reaction (PCR) primers designed with Primer Designer v.2.0 (Scientific & Educational Software, Cary, NC, USA). PCRs were validated using agarose gel electrophoresis. Amplification products for each individual were mixed to create PCR pools, which were purified and quantified. Purification was achieved with a NucleoFast 96 PCR kit (Macherey-Nagel GmbH, Düren, Germany), and quantification of purified amplicons was carried out using an ND-1000 spectrophotometer. The concentration of each PCR pool was then standardized to 0.2 ng/µL. Libraries were prepared with the NexteraXT DNA Library Preparation Kit (Illumina, Inc.) following the manufacturer protocol.

VSX1 gene sequencing analysis was performed using the MiSeq NGS platform (Illumina Inc., San Diego, CA, USA). Concentration and purity of DNA samples were quantified using ND-1000 spectrophotometer (NanoDrop, Wilmington, DE, USA), and the DNA samples were used at 50 ng/µL concentration for analyses. Exons 1-5 of the VSX1 gene and their flanking splice site junctions were amplified using PCR primers designed with Primer Designer v.2.0 (Scientific & Educational Software). PCR products were validated using agarose gel electrophoresis. Additionally, PCR products for each individual were mixed to create PCR pools, which were then purified and quantified. Purifications were done using a NucleoFast ® 96 PCR kit (Macherey-Nagel GmbH & Co. Düren, Germany), and purified PCR products were quantified using ND-1000 spectrophotometer. Quantified PCR pools were standardized to 0.2 ng/µL. The libraries were prepared with the Nextera XT DNA sample preparation kit (Illumina Inc.), following the manufacturer instructions.