Nākamās paaudzes sekvenēšanas metožu izmantošana iedzimtās glaukomas izraisošo ģenētisko faktoru izpētē.
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Date
2023
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
Rīgas Stradiņa universitāte
Rīga Stradiņš University
Rīga Stradiņš University
Abstract
Glaukoma ir iedzimta slimība, kas izraisa neatgriezenisku aklumu. Viens no glaukomas
tipiem ir primāra iedzimta glaukoma, kas parasti attīstās pirmajos gados pēc dzimšanas. Primāra
iedzimta glaukoma var attīstīties arī citu slimību, piemēram, priekšējā segmenta disģenēzes
ietekmē. Ar to ir saistīti vairāki gēni, kā PAX6, JAG1, PATX2, FOXC1,CYP1B1. Nepietiekami
attīstoties priekšējam acs segmentam rodas traucējumi acs šķidruma aizplūdē, palielinot acs
spiedienu, kas ir noteicošais glaukomas attīstībā.
Pētījuma mērķis ir, izmantojot nākamās paaudzes sekvencēšanu, noteikt potenciāli
patogēnus ģenētiskos variantus ar priekšējā segmenta disģenēzi saistītos gēnos, iedzimtas
glaukomas pacientu DNS paraugos.
Pētījumā tika sagatavotas sekvencēšanas bibliotēkas, izmantojot DNBSEQ nanolodīšu
metodi, pacientiem, kuriem ir tikusi diagnosticēta primāra iedzimta glaukoma, kā arī viņu
vecākiem. Iegūtie sekvencēšanas dati tika analizēti, izmantojot Seqr platformu. Iegūto rezultātu
validēšanai tika izmantota Sangera sekvencēšana.
Ar gēnu paneļa sekvencēšanas metodi tika atrasta iedzimta nukleotīdu nomaiņa
c.4357C>A, p.Pro4357Thr PXDN gēnā, kas atrodas 2. hromosomā, vienam pacientam un
vienam no tā vacākiem. Kā arī diviem pacientiem no vienas ģimenes tika identificēta 29
nukleotīdu delēcija BCOR gēnā, kas atrodas X hromosomā. Tā kā nākamās paaudzes
sekvencēšanas dati neļāva precīzi noteikt ģenētisko variantu, bija nepieciešama rezultāta
validēšana ar Sangera sekvencēšanas metodi, kas apstiprināja minēto delēciju.
Glaucoma is a hereditary disease that causes irreversible blindness. One type of glaucoma is primary congenital glaucoma, which usually develops in the first few years after birth. Primary congenital glaucoma can also be caused by other diseases like anterior segment dysgenesis. Several genes such as PAX6, JAG1, PATX2, FOXC1, CYP1B1 are associated with it. Inadequate development of the anterior segment of the eye causes disturbances in the outflow of aqueous humor, increasing the internal eye pressure, which is the determining factor in the development of glaucoma. The aim of the study is to identify potentially pathogenic genetic variants in genes associated with anterior segment dysgenesis in DNA samples from patients with congenital glaucoma using next-generation sequencing. Sequencing libraries in this study were prepared using, the DNBSEQ nanoball method for patients who have been diagnosed with primary congenital glaucoma, as well as their parents. The obtained sequencing data were analyzed using the Seqr platform. Sanger sequencing was used to validate the results. A congenital nucleotide substitution c.4357C>A, p.Pro4357Thr in the PXDN gene, located on chromosome 2, was found by gene panel sequencing method in one patient and one of his parents. Also, two patients from one family were diagnosed with a 29 nucleotide deletion in the BCOR gene, located on the X chromosome. As next - generation sequencing data did not allow precise identification of the genetic variant, it was necessary to use Sanger sequencing for its validation, which confirmed the results obtained by NGS.
Glaucoma is a hereditary disease that causes irreversible blindness. One type of glaucoma is primary congenital glaucoma, which usually develops in the first few years after birth. Primary congenital glaucoma can also be caused by other diseases like anterior segment dysgenesis. Several genes such as PAX6, JAG1, PATX2, FOXC1, CYP1B1 are associated with it. Inadequate development of the anterior segment of the eye causes disturbances in the outflow of aqueous humor, increasing the internal eye pressure, which is the determining factor in the development of glaucoma. The aim of the study is to identify potentially pathogenic genetic variants in genes associated with anterior segment dysgenesis in DNA samples from patients with congenital glaucoma using next-generation sequencing. Sequencing libraries in this study were prepared using, the DNBSEQ nanoball method for patients who have been diagnosed with primary congenital glaucoma, as well as their parents. The obtained sequencing data were analyzed using the Seqr platform. Sanger sequencing was used to validate the results. A congenital nucleotide substitution c.4357C>A, p.Pro4357Thr in the PXDN gene, located on chromosome 2, was found by gene panel sequencing method in one patient and one of his parents. Also, two patients from one family were diagnosed with a 29 nucleotide deletion in the BCOR gene, located on the X chromosome. As next - generation sequencing data did not allow precise identification of the genetic variant, it was necessary to use Sanger sequencing for its validation, which confirmed the results obtained by NGS.
Description
Biomedicīna
Biomedicine
Dzīvās dabas zinātnes
Natural Sciences
Biomedicine
Dzīvās dabas zinātnes
Natural Sciences
Keywords
priekšējā segmenta disģenēze, glaukoma, DNS nanolodīšu sekvencēšanas metode, NGS, anterior segment dysgenesis, glaucoma, DNA nanoball sequencing method, NGS