Semi-nested Taqman real-time quantitative PCR for noninvasive prenatal diagnosis of Bart's hydrops fetalis

Background: Non-invasive prenatal diagnosis based on detection of fetal cell-free DNA is limited when mother and father are both carriers for the same autosomal recessive mutation. Objective: Develop the semi-nested Taqman real-time PCR for quantification of α-thalassemia-1 SEA type deletion allele...

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Main Authors: Sakorn Pornprasert, Kanyakan Sukunthamala, Naowarat Kunyanone, Sririchai Sittiprasert, Khanungnit Thungkham, Sumeth Junorse, Khachonsilp Pongsawatkul, Wisut Pattanaporn, Chantip Jitwong
Format: Journal
Published: 2018
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http://cmuir.cmu.ac.th/jspui/handle/6653943832/51969
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Institution: Chiang Mai University
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spelling th-cmuir.6653943832-519692018-09-04T06:12:42Z Semi-nested Taqman real-time quantitative PCR for noninvasive prenatal diagnosis of Bart's hydrops fetalis Sakorn Pornprasert Kanyakan Sukunthamala Naowarat Kunyanone Sririchai Sittiprasert Khanungnit Thungkham Sumeth Junorse Khachonsilp Pongsawatkul Wisut Pattanaporn Chantip Jitwong Medicine Background: Non-invasive prenatal diagnosis based on detection of fetal cell-free DNA is limited when mother and father are both carriers for the same autosomal recessive mutation. Objective: Develop the semi-nested Taqman real-time PCR for quantification of α-thalassemia-1 SEA type deletion allele in plasma of α-thalassemia-1 SEA carriage pregnancies. Material and Method: Plasma DNA was extracted from six women who carried fetuses with normal, 11 with heterozygote α-thalassemia-1 SEA type deletion and seven with Bart's hydrops fetalis. DNA was amplified using conventional PCR with the primary specific primer set for α-thalassemia-1 SEA type deletion. PCR product was then subjected to the semi-nested realtime PCR using the secondary specific primer and Taqman probe set for α-thalassemia-1 SEA type deletion. The standard curve was constructed using ten-fold serial dilutions of conventional PCR product of the heterozygote α-thalassemia-1 SEA type deletion. Results: Women who carried fetuses with Bart's hydrops fetalis displayed a trend toward higher mean copy number of α-thalassemia-1 SEA type deletion allele vs. women who carried fetuses with normal and heterozygote, albeit not reaching statistical significance. Conclusion: The maternally inherited fetal allele present in maternal plasma is difficult to discern the fetal cell-free DNA from a higher background DNA of the mother. Thus, further investigation is needed to improve the diagnosis of Bart's hydrops fetalis using this technique. 2018-09-04T06:12:42Z 2018-09-04T06:12:42Z 2012-01-01 Journal 01252208 2-s2.0-84856878134 https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=84856878134&origin=inward http://cmuir.cmu.ac.th/jspui/handle/6653943832/51969
institution Chiang Mai University
building Chiang Mai University Library
country Thailand
collection CMU Intellectual Repository
topic Medicine
spellingShingle Medicine
Sakorn Pornprasert
Kanyakan Sukunthamala
Naowarat Kunyanone
Sririchai Sittiprasert
Khanungnit Thungkham
Sumeth Junorse
Khachonsilp Pongsawatkul
Wisut Pattanaporn
Chantip Jitwong
Semi-nested Taqman real-time quantitative PCR for noninvasive prenatal diagnosis of Bart's hydrops fetalis
description Background: Non-invasive prenatal diagnosis based on detection of fetal cell-free DNA is limited when mother and father are both carriers for the same autosomal recessive mutation. Objective: Develop the semi-nested Taqman real-time PCR for quantification of α-thalassemia-1 SEA type deletion allele in plasma of α-thalassemia-1 SEA carriage pregnancies. Material and Method: Plasma DNA was extracted from six women who carried fetuses with normal, 11 with heterozygote α-thalassemia-1 SEA type deletion and seven with Bart's hydrops fetalis. DNA was amplified using conventional PCR with the primary specific primer set for α-thalassemia-1 SEA type deletion. PCR product was then subjected to the semi-nested realtime PCR using the secondary specific primer and Taqman probe set for α-thalassemia-1 SEA type deletion. The standard curve was constructed using ten-fold serial dilutions of conventional PCR product of the heterozygote α-thalassemia-1 SEA type deletion. Results: Women who carried fetuses with Bart's hydrops fetalis displayed a trend toward higher mean copy number of α-thalassemia-1 SEA type deletion allele vs. women who carried fetuses with normal and heterozygote, albeit not reaching statistical significance. Conclusion: The maternally inherited fetal allele present in maternal plasma is difficult to discern the fetal cell-free DNA from a higher background DNA of the mother. Thus, further investigation is needed to improve the diagnosis of Bart's hydrops fetalis using this technique.
format Journal
author Sakorn Pornprasert
Kanyakan Sukunthamala
Naowarat Kunyanone
Sririchai Sittiprasert
Khanungnit Thungkham
Sumeth Junorse
Khachonsilp Pongsawatkul
Wisut Pattanaporn
Chantip Jitwong
author_facet Sakorn Pornprasert
Kanyakan Sukunthamala
Naowarat Kunyanone
Sririchai Sittiprasert
Khanungnit Thungkham
Sumeth Junorse
Khachonsilp Pongsawatkul
Wisut Pattanaporn
Chantip Jitwong
author_sort Sakorn Pornprasert
title Semi-nested Taqman real-time quantitative PCR for noninvasive prenatal diagnosis of Bart's hydrops fetalis
title_short Semi-nested Taqman real-time quantitative PCR for noninvasive prenatal diagnosis of Bart's hydrops fetalis
title_full Semi-nested Taqman real-time quantitative PCR for noninvasive prenatal diagnosis of Bart's hydrops fetalis
title_fullStr Semi-nested Taqman real-time quantitative PCR for noninvasive prenatal diagnosis of Bart's hydrops fetalis
title_full_unstemmed Semi-nested Taqman real-time quantitative PCR for noninvasive prenatal diagnosis of Bart's hydrops fetalis
title_sort semi-nested taqman real-time quantitative pcr for noninvasive prenatal diagnosis of bart's hydrops fetalis
publishDate 2018
url https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=84856878134&origin=inward
http://cmuir.cmu.ac.th/jspui/handle/6653943832/51969
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