Direct detection of alpha satellite DNA with single-base resolution by using abasic Peptide Nucleic Acids and Fluorescent in situ Hybridization.

Autor: Robles-Remacho A; GENYO, Centre for Genomics and Oncological Research, Pfizer, University of Granada, Andalusian Regional Government, PTS Granada, Avenida de la Ilustración, 114, 18016, Granada, Spain; Department of Medicinal and Organic Chemistry, School of Pharmacy, University of Granada, Campus Cartuja s/n, 18071, Granada, Spain; Biosanitary Research Institute of Granada (ibs.GRANADA), University Hospital of Granada/University of Granada, Avenida del Conocimiento, s/n, 18016, Granada, Spain., Luque-Gonzalez MA; GENYO, Centre for Genomics and Oncological Research, Pfizer, University of Granada, Andalusian Regional Government, PTS Granada, Avenida de la Ilustración, 114, 18016, Granada, Spain; Department of Medicinal and Organic Chemistry, School of Pharmacy, University of Granada, Campus Cartuja s/n, 18071, Granada, Spain; Biosanitary Research Institute of Granada (ibs.GRANADA), University Hospital of Granada/University of Granada, Avenida del Conocimiento, s/n, 18016, Granada, Spain., López-Delgado FJ; DESTINA Genomica S.L, PTS Granada, Avenida de la Innovación 1, Edificio BIC, Armilla, 18100, Granada, Spain., Guardia-Monteagudo JJ; DESTINA Genomica S.L, PTS Granada, Avenida de la Innovación 1, Edificio BIC, Armilla, 18100, Granada, Spain., Fara MA; DESTINA Genomica S.L, PTS Granada, Avenida de la Innovación 1, Edificio BIC, Armilla, 18100, Granada, Spain., Pernagallo S; DESTINA Genomica S.L, PTS Granada, Avenida de la Innovación 1, Edificio BIC, Armilla, 18100, Granada, Spain., Sanchez-Martin RM; GENYO, Centre for Genomics and Oncological Research, Pfizer, University of Granada, Andalusian Regional Government, PTS Granada, Avenida de la Ilustración, 114, 18016, Granada, Spain; Department of Medicinal and Organic Chemistry, School of Pharmacy, University of Granada, Campus Cartuja s/n, 18071, Granada, Spain; Biosanitary Research Institute of Granada (ibs.GRANADA), University Hospital of Granada/University of Granada, Avenida del Conocimiento, s/n, 18016, Granada, Spain. Electronic address: rmsanchez@go.ugr.es., Diaz-Mochon JJ; GENYO, Centre for Genomics and Oncological Research, Pfizer, University of Granada, Andalusian Regional Government, PTS Granada, Avenida de la Ilustración, 114, 18016, Granada, Spain; Department of Medicinal and Organic Chemistry, School of Pharmacy, University of Granada, Campus Cartuja s/n, 18071, Granada, Spain; Biosanitary Research Institute of Granada (ibs.GRANADA), University Hospital of Granada/University of Granada, Avenida del Conocimiento, s/n, 18016, Granada, Spain. Electronic address: juanjose.diaz@genyo.es.
Jazyk: angličtina
Zdroj: Biosensors & bioelectronics [Biosens Bioelectron] 2023 Jan 01; Vol. 219, pp. 114770. Date of Electronic Publication: 2022 Oct 07.
DOI: 10.1016/j.bios.2022.114770
Abstrakt: The detection of repetitive sequences with single-base resolution is becoming increasingly important aiming to understand the biological implications of genomic variation in these sequences. However, there is a lack of techniques to experimentally validate sequencing data from repetitive sequences obtained by Next-Generation Sequencing methods, especially in the case of Single-Nucleotide Variations (SNVs). That is one of the reasons why repetitive sequences have been poorly studied and excluded from most genomic studies. Therefore, in addition to sequencing data, there is an urgent need for efficient validation methods of genomic variation in these sequences. Herein we report the development of chemFISH, an alternative method for the detection of SNVs in repetitive sequences. ChemFISH is an innovative method based on dynamic chemistry labelling and abasic Peptide Nucleic Acid (PNA) probes to detect in situ the α-satellite DNA, organized in tandem repeats, with single-base resolution in a direct and rapid reaction. With this approach, we detected by microscopy the α-satellite DNA in a variety of human cell lines, we quantified the detection showing a low coefficient of variation among samples (13.16%-25.33%) and we detected single-base specificity with high sensitivity (82.41%-88.82%). These results indicate that chemFISH can serve as a rapid method to validate previously detected SNVs in sequencing data, as well as to find novel SNVs in repetitive sequences. Furthermore, the versatile chemistry behind chemFISH can lead to develop novel molecular assays for the in situ detection of nucleic acids.
Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Juan Jose Diaz-Mochon reports a relationship with Destina Genomica SL that includes: equity or stocks. Salvatore Pernagallo reports a relationship with Destina Genomica SL that includes: employment and equity or stocks. Francisco Javier Lopez-Delgado reports a relationship with Destina Genomica SL that includes: employment. Juan Jose Guardia-Monteagudo reports a relationship with Destina Genomica SL that includes: employment. Mario Antonio Fara reports a relationship with Destina Genomica SL that includes: employment.
(Copyright © 2022 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
Externí odkaz: