RESUMEN
BACKGROUND: Noninvasive prenatal detection of common fetal aneuploidies with cell-free DNA from maternal plasma has been achieved with high-throughput next-generation sequencing platforms. Turnaround times for previously tested platforms are still unsatisfactory for clinical applications, however, because of the time spent on sequencing. The development of semiconductor sequencing technology has provided a way to shorten overall run times. We studied the feasibility of using semiconductor sequencing technology for the noninvasive detection of fetal aneuploidy. METHODS: Maternal plasma DNA from 13 pregnant women, corresponding to 4 euploid, 6 trisomy 21 (T21), 2 trisomy 18 (T18), and 1 trisomy 13 (T13) pregnancies, were sequenced on the Ion Torrent Personal Genome Machine sequencer platform with 318 chips. The data were analyzed with the T statistic method after correcting for GC bias, and the T value was calculated as an indicator of fetal aneuploidy. RESULTS: We obtained a mean of 3 524 401 high-quality reads per sample, with an efficiency rate of 77.9%. All of the T21, T13, and T18 fetuses could be clearly distinguished from euploid fetuses, and the time spent on library preparation and sequencing was 24 h. CONCLUSIONS: Semiconductor sequencing represents a suitable technology for the noninvasive prenatal detection of fetal aneuploidy. With this platform, sequencing times can be substantially reduced; however, a further larger-scale study is needed to determine the imprecision of noninvasive fetal aneuploidy detection with this system.
Asunto(s)
Trastornos de los Cromosomas/sangre , ADN/química , Feto/patología , Pruebas de Detección del Suero Materno/métodos , Semiconductores , Análisis de Secuencia de ADN/métodos , Trisomía/genética , Trastornos de los Cromosomas/embriología , Cromosomas Humanos Par 13/genética , Cromosomas Humanos Par 18/genética , Cromosomas Humanos Par 21/genética , ADN/sangre , ADN/genética , Estudios de Factibilidad , Femenino , Humanos , Pruebas de Detección del Suero Materno/instrumentación , Embarazo , Análisis de Secuencia de ADN/instrumentación , Trisomía/patologíaRESUMEN
OBJECTIVES: We developed a new human papillomavirus (HPV) genotyping assay based on multiplex polymerase chain reaction and next-generation sequencing (NGS) methods for large-scale cervical cancer screening. METHODS: We first trained the assay on 1,170 self-collected samples, balancing the cutoff points for high-risk types. Then using 4,262 separate self-collected specimens, we compared concordance, sensitivity, and specificity for cervical intraepithelial neoplasia type 2 (CIN2) or higher and CIN type 3 (CIN3) or higher of the HPV sequencing assay with that of Hybrid Capture 2 (HC2) direct samples and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry assay self-samples. RESULTS: All assays had a good agreement. The sensitivity for CIN2 or higher and CIN3 or higher of the self-sampling specimens tested with the sequencing assay run on both MiSeq and Ion Torrent Personal Genome Machine sequencer was similar to that of direct-sampling specimens tested with HC2 (P > .05), but the specificity of the sequencing assay for CIN2 or higher and CIN3 or higher was significantly higher than that of HC2 (P < .01). CONCLUSIONS: This population-based study has demonstrated the applicability of a new NGS high-risk HPV assay for primary cervical cancer screening based on self-collection.
Asunto(s)
Alphapapillomavirus/aislamiento & purificación , Carcinoma de Células Escamosas/virología , Técnicas de Genotipaje/normas , Infecciones por Papillomavirus/virología , Displasia del Cuello del Útero/virología , Neoplasias del Cuello Uterino/virología , Alphapapillomavirus/clasificación , Alphapapillomavirus/genética , Carcinoma de Células Escamosas/diagnóstico , ADN Viral/química , ADN Viral/genética , Detección Precoz del Cáncer , Femenino , Secuenciación de Nucleótidos de Alto Rendimiento , Pruebas de ADN del Papillomavirus Humano , Humanos , Reacción en Cadena de la Polimerasa Multiplex , Infecciones por Papillomavirus/complicaciones , Infecciones por Papillomavirus/diagnóstico , Curva ROC , Sensibilidad y Especificidad , Manejo de Especímenes/métodos , Manejo de Especímenes/normas , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Neoplasias del Cuello Uterino/diagnóstico , Frotis Vaginal , Displasia del Cuello del Útero/diagnósticoRESUMEN
To investigate genetic mechanisms of high altitude adaptations of animals living in the Tibetan Plateau, three mitochondrial genomes (mt-genome) of Tibetan horses living in Naqu (4,500 m) of Tibetan, Zhongdian (3,300 m) and Deqin (3,100 m) of Yunnan province were sequenced. The structures and lengths of these three mt-genomes are similar to the Cheju horse, which is related to Tibetan horses, but little shorter than the Swedish horse. The pair-wise identity of these three horses on nucleotide level is more than 99.3%. When the gene encoding the mitochondrial protein of Tibetan horses was analyzed, we found that NADH6 has higher non-synonymous mutation rate in all of three Tibetan horses. This implies that NADH6 may play a role in Tibetan horses' high altitude adaptation. NADH6 is one of the subunits of the complex I in the respiratory chain. Furthermore, 7 D-loop sequences of Tibetan horse from different areas were sequenced, and the phylogeny tree was constructed to study the origin and evolutionary history of Tibetan horses. The result showed that the genetic diverse was high among Tibetan horses. All of Tibetan horses from Naqu were clustered into one clade, and Tibetan horses from Zhongdian and Deqin were clustered into others clades. The first molecular evidence of Tibetan horses indicated in this study is that Tibetan horse population might have multiple origins.