Application of single sperm sequencing combined with PCR-reverse dot blot in preimplantation genetic testing of Southeast Asian deletion type α thalassemia
10.3760/cma.j.cn101441-20220104-00002
- VernacularTitle:单精子测序结合PCR-反向点杂交技术在1例东南亚缺失型α地中海贫血患者胚胎植入前遗传学检测中的应用
- Author:
Jianchun HE
1
;
Qing LI
1
;
Yanchao WANG
1
;
Jiajia XIAN
1
;
Mincong ZHANG
1
;
Wenzhi HE
1
;
Xiaoyan MA
1
;
Guoxin YE
1
;
Xiaoman WANG
1
;
Shaoying LI
1
Author Information
1. 广州医科大学附属第三医院妇产科 广东省产科重大疾病重点实验室 妇产科研究所实验部，广州 510150
- Publication Type:Journal Article
- Keywords:
Genetic linkage;
Polymorphism, single nucleotide;
Preimplantation diagnosis;
Single sperm sequencing;
Southeast Asian deletion type α thalassemia
- From:
Chinese Journal of Reproduction and Contraception
2023;43(4):407-411
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the value of single sperm sequencing combined with PCR-reverse dot blot (PCR-RDB) technology in preimplantation genetic testing (PGT) of Southeast Asian deletion type α thalassemia.Methods:A couple of Southeast Asian deletion type α thalassemia carrier was selected in this case, who asked for assisted reproduction in the Department of Obstetrics and Gynecology of the Third Affiliated Hospital of Guangzhou Medical University on April 24, 2020. The male carrier was subjected to single sperm isolation by the swim-up method combined with micropipette due to his incomplete pedigree. Five single sperm samples were isolated and their whole-genome were amplified. The genotype of thalassemia of single sperm samples were determined by PCR-RDB. SNPs were selected as genetic markers in the range of 1 Mb upstream and downstream of -- SEA region to construct chromosomal haplotype. Next, we performed whole-genome amplification on six blastocyst biopsy samples, and then next-generation sequencing was carried out to detect the carrier status of the embryos. Finally, the non-pathogenic blastocysts were selected for transplantation, and the preimplantation genetic testing for monogenic (PGT-M) results were confirmed by the prenatal genetic diagnosis during pregnancy. Results:The pathogenic variant in the female was inherited from her mother, and four of the five single sperm samples were wild-type. Ten male informative SNP markers were identified by single sperm sequencing, and six female informative SNP markers were identified by linkage analysis of female family. PGT-M results indicated that four blastocysts were αα/-- SEA and two were -- SEA/-- SEA. The result of prenatal diagnosis revealed that the fetus was a carrier of -- SEA , which was consistent with PGT-M result, and a healthy girl was delivered at 40 weeks of gestation. Conclusion:For male carriers of Southeast Asian deletion type α thalassemia with incomplete pedigree, single sperm sequencing combined with PCR-RDB technology can be used to select SNP sites, and PGT-M can be performed by linkage analysis.
