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Amplicon Sequencing-Based Noninvasive Fetal Genotyping for RHD-Positive D Antigen-Negative Alleles
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Amplicon Sequencing-Based Noninvasive Fetal Genotyping for RHD-Positive D Antigen-Negative Alleles
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Journal Article
Amplicon Sequencing-Based Noninvasive Fetal Genotyping for RHD-Positive D Antigen-Negative Alleles
2019
Overview
To avoid hemolytic disease of the fetus and newborn resulting from maternal alloantibodies against fetal Rh antigens, anti-D immunoglobulin is routinely administered to RhD-negative pregnant women in Japan. Fetal
genotyping using cell-free DNA may prevent unnecessary antibody administration; however, current PCR-based methods, which detect
deletion, do not address the higher rates of
-positive D antigen-negative alleles in nonwhite populations without additional inspections.
We developed an amplicon-sequencing method that could estimate the type of paternally inherited fetal
allele from 4 major
alleles in the Japanese population: the D antigen-positive allele (
*
, 92.9%) and 3 D antigen-negative alleles (
*
, 6.6%;
*
, 0.3%;
*
, 0.1%) using cell-free DNA obtained from the blood plasma of pregnant women.
The method correctly determined the fetal RhD type even when RhD-negative pregnant women possessed an
-positive D antigen-negative allele:
*
or
*
.
This method is a reliable noninvasive fetal
genotyping method for Japanese and other East Asian populations. The genotyping principle of amplifying 2 different regions using the same primer pair and distinguishing them by their sequence difference during the subsequent mapping procedure is also theoretically applicable to
-positive D antigen-negative alleles prevalent in Africans. Therefore, this method offers an opportunity to consider targeted administration of anti-D immunoglobulin to RhD-negative pregnant women in East Asian and African countries and to increase the specificity of the fetal
genotyping implemented nationwide in several European countries.
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