Targeted next-generation sequencing and long-read HiFi sequencing provide novel insights into clinically significant KLF1 variants
Abstract Background Krüppel-like factor 1 (KLF1), a crucial erythroid transcription factor, plays a significant role in various erythroid changes and haemolytic diseases. The rare erythrocyte Lutheran inhibitor (In(Lu)) blood group phenotype serves as an effective model for identifying KLF1 hypomorp...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Language: | English |
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BMC
2024-03-01
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| Series: | BMC Genomics |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s12864-024-10148-x |
| _version_ | 1827328547688022016 |
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| author | Luyi Ye Chen Wang Aijing Li Minghao Li Yan Pi Jingmin Yang Ziyan Zhu Daru Lu |
| author_facet | Luyi Ye Chen Wang Aijing Li Minghao Li Yan Pi Jingmin Yang Ziyan Zhu Daru Lu |
| author_sort | Luyi Ye |
| collection | DOAJ |
| description | Abstract Background Krüppel-like factor 1 (KLF1), a crucial erythroid transcription factor, plays a significant role in various erythroid changes and haemolytic diseases. The rare erythrocyte Lutheran inhibitor (In(Lu)) blood group phenotype serves as an effective model for identifying KLF1 hypomorphic and loss-of-function variants. In this study, we aimed to analyse the genetic background of the In(Lu) phenotype in a population-based sample group by high-throughput technologies to find potentially clinically significant KLF1 variants. Results We included 62 samples with In(Lu) phenotype, screened from over 300,000 Chinese blood donors. Among them, 36 samples were sequenced using targeted Next Generation Sequencing (NGS), whereas 19 samples were sequenced using High Fidelity (HiFi) technology. In addition, seven samples were simply sequenced using Sanger sequencing. A total of 29 hypomorphic or loss-of-function variants of KLF1 were identified, 21 of which were newly discovered. All new variants discovered by targeted NGS or HiFi sequencing were validated through Sanger sequencing, and the obtained results were found to be consistent. The KLF1 haplotypes of all new variants were further confirmed using clone sequencing or HiFi sequencing. The lack of functional KLF1 variants detected in the four samples indicates the presence of additional regulatory mechanisms. In addition, some samples exhibited BCAM polymorphisms, which encodes antigens of the Lutheran (LU) blood group system. However, no BCAM mutations which leads to the absence of LU proteins were detected. Conclusions High-throughput sequencing methods, particularly HiFi sequencing, were introduced for the first time into genetic analysis of the In(Lu) phenotype. Targeted NGS and HiFi sequencing demonstrated the accuracy of the results, providing additional advantages such as simultaneous analysis of other blood group genes and clarification of haplotypes. Using the In(Lu) phenotype, a powerful model for identifying hypomorphic or loss-of-function KLF1 variants, numerous novel variants have been detected, which have contributed to the comprehensive understanding of KLF1. These clinically significant KLF1 mutations can serve as a valuable reference for the diagnosis of related blood cell diseases. |
| first_indexed | 2024-03-07T15:19:16Z |
| format | Article |
| id | doaj.art-b1d9890802c34b3e9b1dce5b68f0409e |
| institution | Directory Open Access Journal |
| issn | 1471-2164 |
| language | English |
| last_indexed | 2024-03-07T15:19:16Z |
| publishDate | 2024-03-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Genomics |
| spelling | doaj.art-b1d9890802c34b3e9b1dce5b68f0409e2024-03-05T17:46:35ZengBMCBMC Genomics1471-21642024-03-0125111110.1186/s12864-024-10148-xTargeted next-generation sequencing and long-read HiFi sequencing provide novel insights into clinically significant KLF1 variantsLuyi Ye0Chen Wang1Aijing Li2Minghao Li3Yan Pi4Jingmin Yang5Ziyan Zhu6Daru Lu7State Key Laboratory of Genetic Engineering and MOE Engineering Research Center of Gene Technology, School of Life Sciences, Fudan UniversityShanghai Institute of Blood Transfusion, Shanghai Blood CenterShanghai Institute of Blood Transfusion, Shanghai Blood CenterShanghai Institute of Blood Transfusion, Shanghai Blood CenterState Key Laboratory of Genetic Engineering and MOE Engineering Research Center of Gene Technology, School of Life Sciences, Fudan UniversityState Key Laboratory of Genetic Engineering and MOE Engineering Research Center of Gene Technology, School of Life Sciences, Fudan UniversityShanghai Institute of Blood Transfusion, Shanghai Blood CenterState Key Laboratory of Genetic Engineering and MOE Engineering Research Center of Gene Technology, School of Life Sciences, Fudan UniversityAbstract Background Krüppel-like factor 1 (KLF1), a crucial erythroid transcription factor, plays a significant role in various erythroid changes and haemolytic diseases. The rare erythrocyte Lutheran inhibitor (In(Lu)) blood group phenotype serves as an effective model for identifying KLF1 hypomorphic and loss-of-function variants. In this study, we aimed to analyse the genetic background of the In(Lu) phenotype in a population-based sample group by high-throughput technologies to find potentially clinically significant KLF1 variants. Results We included 62 samples with In(Lu) phenotype, screened from over 300,000 Chinese blood donors. Among them, 36 samples were sequenced using targeted Next Generation Sequencing (NGS), whereas 19 samples were sequenced using High Fidelity (HiFi) technology. In addition, seven samples were simply sequenced using Sanger sequencing. A total of 29 hypomorphic or loss-of-function variants of KLF1 were identified, 21 of which were newly discovered. All new variants discovered by targeted NGS or HiFi sequencing were validated through Sanger sequencing, and the obtained results were found to be consistent. The KLF1 haplotypes of all new variants were further confirmed using clone sequencing or HiFi sequencing. The lack of functional KLF1 variants detected in the four samples indicates the presence of additional regulatory mechanisms. In addition, some samples exhibited BCAM polymorphisms, which encodes antigens of the Lutheran (LU) blood group system. However, no BCAM mutations which leads to the absence of LU proteins were detected. Conclusions High-throughput sequencing methods, particularly HiFi sequencing, were introduced for the first time into genetic analysis of the In(Lu) phenotype. Targeted NGS and HiFi sequencing demonstrated the accuracy of the results, providing additional advantages such as simultaneous analysis of other blood group genes and clarification of haplotypes. Using the In(Lu) phenotype, a powerful model for identifying hypomorphic or loss-of-function KLF1 variants, numerous novel variants have been detected, which have contributed to the comprehensive understanding of KLF1. These clinically significant KLF1 mutations can serve as a valuable reference for the diagnosis of related blood cell diseases.https://doi.org/10.1186/s12864-024-10148-xKLF1In(Lu) phenotypeLU blood groupNGSHiFi sequencing |
| spellingShingle | Luyi Ye Chen Wang Aijing Li Minghao Li Yan Pi Jingmin Yang Ziyan Zhu Daru Lu Targeted next-generation sequencing and long-read HiFi sequencing provide novel insights into clinically significant KLF1 variants BMC Genomics KLF1 In(Lu) phenotype LU blood group NGS HiFi sequencing |
| title | Targeted next-generation sequencing and long-read HiFi sequencing provide novel insights into clinically significant KLF1 variants |
| title_full | Targeted next-generation sequencing and long-read HiFi sequencing provide novel insights into clinically significant KLF1 variants |
| title_fullStr | Targeted next-generation sequencing and long-read HiFi sequencing provide novel insights into clinically significant KLF1 variants |
| title_full_unstemmed | Targeted next-generation sequencing and long-read HiFi sequencing provide novel insights into clinically significant KLF1 variants |
| title_short | Targeted next-generation sequencing and long-read HiFi sequencing provide novel insights into clinically significant KLF1 variants |
| title_sort | targeted next generation sequencing and long read hifi sequencing provide novel insights into clinically significant klf1 variants |
| topic | KLF1 In(Lu) phenotype LU blood group NGS HiFi sequencing |
| url | https://doi.org/10.1186/s12864-024-10148-x |
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