Validation of a next-generation sequencing oncology panel optimized for low input DNA.
Cancer Genet
; 228-229: 55-63, 2018 12.
Article
en En
| MEDLINE
| ID: mdl-30553474
ABSTRACT
One caveat of next-generation sequencing (NGS)-based clinical oncology testing is the high amount of input DNA required. We sought to develop a focused NGS panel that could capture hotspot regions in relevant genes requiring 0.5-10â¯ng input DNA. The resulting Penn Precision Panel (PPP) targeted 20 genes containing clinically significant variants relevant to many cancers. One hundred twenty-three samples were analyzed, including 83 solid tumor specimens derived from FFPE. Various input quantities of DNA (0.5-10â¯ng) were amplified with content-specific PCR primer pools, then sequenced on a MiSeq instrument (Illumina, Inc.) via paired-end, 2â¯×â¯186 base pair reads to an average read depth of greater than 6500x. Variants were detected using an in-house analysis pipeline. Clinical sensitivity and specificity were assessed using results from our previously validated solid tumor NGS panel; sensitivity of the PPP is 96.75% (387/400 variants) and specificity is 99.9% (8427/8428 base pairs). Variant allele frequencies (VAFs) are highly concordant across both assays (râ¯=â¯0.98 pâ¯<â¯0.0001). The PPP is a robust, clinically validated test optimized for low-yield solid tumor specimens, capturing a high percentage of clinically relevant variants found by larger commercially available NGS panels while using only 0.5-10â¯ng of input DNA.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
ADN de Neoplasias
/
Secuenciación de Nucleótidos de Alto Rendimiento
Límite:
Humans
Idioma:
En
Revista:
Cancer Genet
Año:
2018
Tipo del documento:
Article
País de afiliación:
Estados Unidos