Coupling of Single Molecule, Long Read Sequencing with IMGT/HighV-QUEST Analysis Expedites Identification of SIV gp140-Specific Antibodies from scFv Phage Display Libraries
The simian immunodeficiency virus (SIV)/macaque model of human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome pathogenesis is critical for furthering our understanding of the role of antibody responses in the prevention of HIV infection, and will only increase in importance as macaq...
| Main Authors: | , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2018-03-01
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| Series: | Frontiers in Immunology |
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| Online Access: | http://journal.frontiersin.org/article/10.3389/fimmu.2018.00329/full |
| _version_ | 1811306948011753472 |
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| author | Seung Yub Han Alesia Antoine David Howard Bryant Chang Woo Sung Chang Matthew Slein Gintaras Deikus Sofia Kossida Patrice Duroux Marie-Paule Lefranc Robert P. Sebra Melissa L. Smith Ismael Ben F. Fofana |
| author_facet | Seung Yub Han Alesia Antoine David Howard Bryant Chang Woo Sung Chang Matthew Slein Gintaras Deikus Sofia Kossida Patrice Duroux Marie-Paule Lefranc Robert P. Sebra Melissa L. Smith Ismael Ben F. Fofana |
| author_sort | Seung Yub Han |
| collection | DOAJ |
| description | The simian immunodeficiency virus (SIV)/macaque model of human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome pathogenesis is critical for furthering our understanding of the role of antibody responses in the prevention of HIV infection, and will only increase in importance as macaque immunoglobulin (IG) gene databases are expanded. We have previously reported the construction of a phage display library from a SIV-infected rhesus macaque (Macaca mulatta) using oligonucleotide primers based on human IG gene sequences. Our previous screening relied on Sanger sequencing, which was inefficient and generated only a few dozen sequences. Here, we re-analyzed this library using single molecule, real-time (SMRT) sequencing on the Pacific Biosciences (PacBio) platform to generate thousands of highly accurate circular consensus sequencing (CCS) reads corresponding to full length single chain fragment variable. CCS data were then analyzed through the international ImMunoGeneTics information system® (IMGT®)/HighV-QUEST (www.imgt.org) to identify variable genes and perform statistical analyses. Overall the library was very diverse, with 2,569 different IMGT clonotypes called for the 5,238 IGHV sequences assigned to an IMGT clonotype. Within the library, SIV-specific antibodies represented a relatively limited number of clones, with only 135 different IMGT clonotypes called from 4,594 IGHV-assigned sequences. Our data did confirm that the IGHV4 and IGHV3 gene usage was the most abundant within the rhesus antibodies screened, and that these genes were even more enriched among SIV gp140-specific antibodies. Although a broad range of VH CDR3 amino acid (AA) lengths was observed in the unpanned library, the vast majority of SIV gp140-specific antibodies demonstrated a more uniform VH CDR3 length (20 AA). This uniformity was far less apparent when VH CDR3 were classified according to their clonotype (range: 9–25 AA), which we believe is more relevant for specific antibody identification. Only 174 IGKV and 588 IGLV clonotypes were identified within the VL sequences associated with SIV gp140-specific VH. Together, these data strongly suggest that the combination of SMRT sequencing with the IMGT/HighV-QUEST querying tool will facilitate and expedite our understanding of polyclonal antibody responses during SIV infection and may serve to rapidly expand the known scope of macaque V genes utilized during these responses. |
| first_indexed | 2024-04-13T08:54:49Z |
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| institution | Directory Open Access Journal |
| issn | 1664-3224 |
| language | English |
| last_indexed | 2024-04-13T08:54:49Z |
| publishDate | 2018-03-01 |
| publisher | Frontiers Media S.A. |
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| series | Frontiers in Immunology |
| spelling | doaj.art-d31146ce90f14af4af6e11d84997c8192022-12-22T02:53:20ZengFrontiers Media S.A.Frontiers in Immunology1664-32242018-03-01910.3389/fimmu.2018.00329313487Coupling of Single Molecule, Long Read Sequencing with IMGT/HighV-QUEST Analysis Expedites Identification of SIV gp140-Specific Antibodies from scFv Phage Display LibrariesSeung Yub Han0Alesia Antoine1David Howard2Bryant Chang3Woo Sung Chang4Matthew Slein5Gintaras Deikus6Sofia Kossida7Patrice Duroux8Marie-Paule Lefranc9Robert P. Sebra10Melissa L. Smith11Ismael Ben F. Fofana12Biology Department, Boston College, Chestnut Hill, MA, United StatesDepartment of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, Icahn Institute of Genomics and Multiscale Biology, New York, NY, United StatesBiology Department, Boston College, Chestnut Hill, MA, United StatesBiology Department, Boston College, Chestnut Hill, MA, United StatesBiology Department, Boston College, Chestnut Hill, MA, United StatesBiology Department, Boston College, Chestnut Hill, MA, United StatesDepartment of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, Icahn Institute of Genomics and Multiscale Biology, New York, NY, United StatesThe international ImMunoGeneTics information system® (IMGT®), Laboratoire d'ImmunoGénétique Moléculaire (LIGM), Institut de Génétique Humaine (IGH), UMR CNRS, Montpellier University, Montpellier, FranceThe international ImMunoGeneTics information system® (IMGT®), Laboratoire d'ImmunoGénétique Moléculaire (LIGM), Institut de Génétique Humaine (IGH), UMR CNRS, Montpellier University, Montpellier, FranceThe international ImMunoGeneTics information system® (IMGT®), Laboratoire d'ImmunoGénétique Moléculaire (LIGM), Institut de Génétique Humaine (IGH), UMR CNRS, Montpellier University, Montpellier, FranceDepartment of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, Icahn Institute of Genomics and Multiscale Biology, New York, NY, United StatesDepartment of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, Icahn Institute of Genomics and Multiscale Biology, New York, NY, United StatesBiology Department, Boston College, Chestnut Hill, MA, United StatesThe simian immunodeficiency virus (SIV)/macaque model of human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome pathogenesis is critical for furthering our understanding of the role of antibody responses in the prevention of HIV infection, and will only increase in importance as macaque immunoglobulin (IG) gene databases are expanded. We have previously reported the construction of a phage display library from a SIV-infected rhesus macaque (Macaca mulatta) using oligonucleotide primers based on human IG gene sequences. Our previous screening relied on Sanger sequencing, which was inefficient and generated only a few dozen sequences. Here, we re-analyzed this library using single molecule, real-time (SMRT) sequencing on the Pacific Biosciences (PacBio) platform to generate thousands of highly accurate circular consensus sequencing (CCS) reads corresponding to full length single chain fragment variable. CCS data were then analyzed through the international ImMunoGeneTics information system® (IMGT®)/HighV-QUEST (www.imgt.org) to identify variable genes and perform statistical analyses. Overall the library was very diverse, with 2,569 different IMGT clonotypes called for the 5,238 IGHV sequences assigned to an IMGT clonotype. Within the library, SIV-specific antibodies represented a relatively limited number of clones, with only 135 different IMGT clonotypes called from 4,594 IGHV-assigned sequences. Our data did confirm that the IGHV4 and IGHV3 gene usage was the most abundant within the rhesus antibodies screened, and that these genes were even more enriched among SIV gp140-specific antibodies. Although a broad range of VH CDR3 amino acid (AA) lengths was observed in the unpanned library, the vast majority of SIV gp140-specific antibodies demonstrated a more uniform VH CDR3 length (20 AA). This uniformity was far less apparent when VH CDR3 were classified according to their clonotype (range: 9–25 AA), which we believe is more relevant for specific antibody identification. Only 174 IGKV and 588 IGLV clonotypes were identified within the VL sequences associated with SIV gp140-specific VH. Together, these data strongly suggest that the combination of SMRT sequencing with the IMGT/HighV-QUEST querying tool will facilitate and expedite our understanding of polyclonal antibody responses during SIV infection and may serve to rapidly expand the known scope of macaque V genes utilized during these responses.http://journal.frontiersin.org/article/10.3389/fimmu.2018.00329/fullantibodysimian immunodeficiency virusrhesus macaquePacBio sequencingsingle chain fragment variable libraryphage display |
| spellingShingle | Seung Yub Han Alesia Antoine David Howard Bryant Chang Woo Sung Chang Matthew Slein Gintaras Deikus Sofia Kossida Patrice Duroux Marie-Paule Lefranc Robert P. Sebra Melissa L. Smith Ismael Ben F. Fofana Coupling of Single Molecule, Long Read Sequencing with IMGT/HighV-QUEST Analysis Expedites Identification of SIV gp140-Specific Antibodies from scFv Phage Display Libraries Frontiers in Immunology antibody simian immunodeficiency virus rhesus macaque PacBio sequencing single chain fragment variable library phage display |
| title | Coupling of Single Molecule, Long Read Sequencing with IMGT/HighV-QUEST Analysis Expedites Identification of SIV gp140-Specific Antibodies from scFv Phage Display Libraries |
| title_full | Coupling of Single Molecule, Long Read Sequencing with IMGT/HighV-QUEST Analysis Expedites Identification of SIV gp140-Specific Antibodies from scFv Phage Display Libraries |
| title_fullStr | Coupling of Single Molecule, Long Read Sequencing with IMGT/HighV-QUEST Analysis Expedites Identification of SIV gp140-Specific Antibodies from scFv Phage Display Libraries |
| title_full_unstemmed | Coupling of Single Molecule, Long Read Sequencing with IMGT/HighV-QUEST Analysis Expedites Identification of SIV gp140-Specific Antibodies from scFv Phage Display Libraries |
| title_short | Coupling of Single Molecule, Long Read Sequencing with IMGT/HighV-QUEST Analysis Expedites Identification of SIV gp140-Specific Antibodies from scFv Phage Display Libraries |
| title_sort | coupling of single molecule long read sequencing with imgt highv quest analysis expedites identification of siv gp140 specific antibodies from scfv phage display libraries |
| topic | antibody simian immunodeficiency virus rhesus macaque PacBio sequencing single chain fragment variable library phage display |
| url | http://journal.frontiersin.org/article/10.3389/fimmu.2018.00329/full |
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