Unveiling APOL1 Haplotypes : A Novel Classification Through Probe-Independent Quantitative Real-Time PCR
Introduction: Apolipoprotein-L1 (APOL1) is a primate-specific protein component of high- density lipoprotein (HDL). Two variants of APOL1 (G1 and G2), provide resistance to parasitic infections in African Americans but are also implicated in kidney-related diseases and transplant outcomes in recipients. This study aims to identify these risk variants using a novel probe- independent quantitative real-time PCR method in a high African American recipient cohort. Additionally, it aims to develop a new stratification approach based on haplotype-centric model.
Methods: Genomic DNA was extracted from recipient PBMCs using SDS lysis buffer and proteinase K. Quantitative PCR assay with modified forward primers and a common reverse primer enabled us to identify single nucleotide polymorphisms (SNPs) and the 6-bp deletion quantitatively. Additionally, we used sanger sequencing to verify our QPCR findings.
Results: Our novel probe-independent qPCR effectively distinguished homozygous wild-type, heterozygous SNPs/deletion, and homozygous SNPs/deletion, with at least 4-fold differences. High prevalence of APOL1 variants was observed (18% two-risk alleles, 34% one-risk allele) in our recipient cohort. Intriguingly, up to 12-month follow-up revealed no significant impact of recipient APOL1 variants on transplant outcomes. Ongoing research will encompass more time points and a larger patient cohort, allowing a comprehensive evaluation of G1/G2 variant subgroups categorized by new haplotype scores, enriching our understanding.
Conclusions: Our cost-effective and rapid qPCR technique facilitates APOL1 genotyping within hours. Prospective and retrospective studies will enable comparisons with long-term allograft rejection, potentially predicting early/late-stage transplant outcomes based on haplotype evaluation in this diverse group of kidney transplant recipients.
Medienart: |
E-Artikel |
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Erscheinungsjahr: |
2023 |
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Erschienen: |
2023 |
Enthalten in: |
Zur Gesamtaufnahme - year:2023 |
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Enthalten in: |
bioRxiv : the preprint server for biology - (2023) vom: 19. Okt. |
Sprache: |
Englisch |
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Beteiligte Personen: |
Dogan, Murat [VerfasserIn] |
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Anmerkungen: |
Date Revised 31.10.2023 published: Electronic Citation Status PubMed-not-MEDLINE |
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doi: |
10.1101/2023.10.16.562539 |
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funding: |
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PPN (Katalog-ID): |
NLM363970495 |
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520 | |a Introduction: Apolipoprotein-L1 (APOL1) is a primate-specific protein component of high- density lipoprotein (HDL). Two variants of APOL1 (G1 and G2), provide resistance to parasitic infections in African Americans but are also implicated in kidney-related diseases and transplant outcomes in recipients. This study aims to identify these risk variants using a novel probe- independent quantitative real-time PCR method in a high African American recipient cohort. Additionally, it aims to develop a new stratification approach based on haplotype-centric model | ||
520 | |a Methods: Genomic DNA was extracted from recipient PBMCs using SDS lysis buffer and proteinase K. Quantitative PCR assay with modified forward primers and a common reverse primer enabled us to identify single nucleotide polymorphisms (SNPs) and the 6-bp deletion quantitatively. Additionally, we used sanger sequencing to verify our QPCR findings | ||
520 | |a Results: Our novel probe-independent qPCR effectively distinguished homozygous wild-type, heterozygous SNPs/deletion, and homozygous SNPs/deletion, with at least 4-fold differences. High prevalence of APOL1 variants was observed (18% two-risk alleles, 34% one-risk allele) in our recipient cohort. Intriguingly, up to 12-month follow-up revealed no significant impact of recipient APOL1 variants on transplant outcomes. Ongoing research will encompass more time points and a larger patient cohort, allowing a comprehensive evaluation of G1/G2 variant subgroups categorized by new haplotype scores, enriching our understanding | ||
520 | |a Conclusions: Our cost-effective and rapid qPCR technique facilitates APOL1 genotyping within hours. Prospective and retrospective studies will enable comparisons with long-term allograft rejection, potentially predicting early/late-stage transplant outcomes based on haplotype evaluation in this diverse group of kidney transplant recipients | ||
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700 | 1 | |a Watkins, Christine |e verfasserin |4 aut | |
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700 | 1 | |a Gower, Elizabeth G |e verfasserin |4 aut | |
700 | 1 | |a Eason, James D |e verfasserin |4 aut | |
700 | 1 | |a Bhalla, Anshul |e verfasserin |4 aut | |
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700 | 1 | |a Nezakatgoo, Nosratollah |e verfasserin |4 aut | |
700 | 1 | |a Eymard, Corey |e verfasserin |4 aut | |
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700 | 1 | |a Bajwa, Amandeep |e verfasserin |4 aut | |
700 | 1 | |a Kuscu, Canan |e verfasserin |4 aut | |
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