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Phagemid
Phagemid
Phagemid
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A phagemid or phasmid is a DNA-based cloning vector, which has both bacteriophage and plasmid properties.[1] These vectors carry, in addition to the origin of plasmid replication, an origin of replication derived from bacteriophage. Unlike commonly used plasmids, phagemid vectors differ by having the ability to be packaged into the capsid of a bacteriophage, due to their having a genetic sequence that signals for packaging. Phagemids are used in a variety of biotechnology applications; for example, they can be used in a molecular biology technique called "phage display".[2]

The term "phagemid" or "phagemids" was coined by a group of Soviet scientists, who discovered them, named them, and published the article in April 1984 in Gene magazine.[3]

Properties of the cloning vector

[edit]

A phagemid (plasmid + phage) is a plasmid that contains an f1 origin of replication from an f1 phage.[4] It can be used as a type of cloning vector in combination with filamentous phage M13. A phagemid can be replicated as a plasmid, and also be packaged as single stranded DNA in viral particles. Phagemids contain an origin of replication (ori) for double stranded replication, as well as an f1 ori to enable single stranded replication and packaging into phage particles.[4] Many commonly used plasmids contain an f1 ori and are thus phagemids.

Similarly to a plasmid, a phagemid can be used to clone DNA fragments and be introduced into a bacterial host by a range of techniques, such as transformation and electroporation. However, infection of a bacterial host containing a phagemid with a 'helper' phage, for example VCSM13 or M13K07, provides the necessary viral components to enable single stranded DNA replication and packaging of the phagemid DNA into phage particles. The 'helper' phage infects the bacterial host by first attaching to the host cell's pilus and then, after attachment, transporting the phage genome into the cytoplasm of the host cell. Inside the cell, the phage genome triggers production of single stranded phagemid DNA in the cytoplasm. This phagemid DNA is then packaged into phage particles. The phage particles containing ssDNA are released from the bacterial host cell into the extracellular environment.

Filamentous phages retard bacterial growth but, contrasting with the lambda phage and the T7 phage, are not generally lytic. Helper phages are usually engineered to package less efficiently (via a defective phage origin of replication)[5] than the phagemid so that the resultant phage particles contain predominantly phagemid DNA. F1 Filamentous phage infection requires the presence of a pilus so only bacterial hosts containing the F-plasmid or its derivatives can be used to generate phage particles.

Prior to the development of cycle sequencing, phagemids were used to generate single stranded DNA template for sequencing purposes. Today phagemids are still useful for generating templates for site-directed mutagenesis. Detailed characterisation of the filamentous phage life cycle and structural features lead to the development of phage display technology, in which a range of peptides and proteins can be expressed as fusions to phage coat proteins and displayed on the viral surface. The displayed peptides and polypeptides are associated with the corresponding coding DNA within the phage particle and so this technique lends itself to the study of protein-protein interactions and other ligand/receptor combinations.

References

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Phagemid

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from Grokipedia
A phagemid is a hybrid in that merges the replicative properties of a with specific genetic elements from filamentous bacteriophages, such as the M13 phage, enabling both double-stranded DNA propagation in bacterial hosts like and the production of single-stranded DNA packaged into infectious phage-like particles upon with a helper phage. These vectors typically include a plasmid origin of replication (e.g., ), an resistance for selection, the phage f1 origin for single-stranded DNA synthesis, and often a fusion to one phage coat protein (such as gIII encoding the minor coat protein pIII), but lack the full set of phage structural genes, necessitating the helper phage to provide essential proteins for particle assembly. This design allows phagemids to achieve higher transformation efficiencies and larger library sizes compared to full phage vectors, while maintaining stability under diverse conditions like extreme pH or temperature. Phagemids were developed in the 1980s, building on the foundational technique invented by George P. Smith in 1985, with early examples including constructs for filamentous phages like M13. The primary application of phagemids lies in phage display technology, where they facilitate the construction of combinatorial libraries expressing diverse peptides, antibody fragments (e.g., scFv or Fab), or proteins fused to the pIII coat protein, enabling affinity-based selection (biopanning) to isolate high-affinity binders against targets like antigens or small molecules. Unlike multivalent display on full phage vectors, phagemids typically enable monovalent or controlled-valency display by relying on wild-type coat proteins from the helper phage, which preserves particle and avoids effects that can bias selection toward low-affinity clones. This has revolutionized fields such as antibody engineering, vaccine development, and protein therapeutics, with phagemid-derived libraries routinely yielding candidates for clinical use, including monoclonal antibodies approved for cancer and infectious disease treatments. Beyond display, phagemids support applications in , , and targeted , such as engineering antibacterial particles that produce toxins upon infection.

Definition and Properties

Definition

A phagemid is a DNA-based that combines the replicative and structural properties of both and bacteriophages, enabling efficient propagation of inserted genetic material in bacterial hosts. These hybrid vectors typically derive from filamentous phages like M13, incorporating a for stable maintenance as double-stranded DNA in , while lacking most phage genes necessary for independent virion assembly. At its core, the hybrid nature of a phagemid allows it to replicate autonomously as a plasmid within bacteria but to be packaged into phage-like particles (virions) when co-infected with a helper phage that supplies essential coat proteins. This packaging mechanism facilitates the production of single-stranded DNA, a key feature for applications in sequencing and display technologies. Phagemids play a central role in molecular biology techniques, such as gene cloning and protein engineering.

Key Properties

Phagemids exhibit a unique hybrid functionality that allows them to replicate as double-stranded DNA (dsDNA) within bacterial hosts, relying on a plasmid origin of replication such as ColE1 or p15A for stable maintenance and propagation as multicopy plasmids. This dsDNA replication mode enables efficient cloning and amplification in Escherichia coli without the need for additional viral components. A distinguishing feature is their capacity to generate single-stranded DNA (ssDNA) through the incorporation of phage-derived packaging signals, including the f1 from M13, which initiate the production of ssDNA upon induction. However, phagemids lack the genes encoding phage coat proteins, necessitating co-infection with a helper phage—typically M13 derivatives like VCSM13 or M13KO7—to supply these proteins in trans for the ssDNA into filamentous virions. This dependency on helper phages ensures controlled ssDNA production while minimizing the risk of uncontrolled phage propagation. Phagemids generally range from 3 to 10 kilobases in size, accommodating inserts while maintaining a compact backbone for ease of manipulation. They commonly incorporate selectable markers, such as antibiotic resistance genes for or kanamycin, to allow for the selection and maintenance of transformed bacterial cells.

History

Discovery

Phagemids, as hybrid vectors combining plasmid replication with elements from filamentous bacteriophages like M13 or f1, were first developed in 1983 with the pEMBL series of plasmids by Dente, Cesareni, and . These vectors incorporated a origin (e.g., pBR322-derived), an antibiotic resistance marker, and the f1 phage origin of replication, enabling double-stranded propagation in and single-stranded DNA production upon with helper phage. This built on 1970s advances in and early 1980s work on filamentous phage derivatives for genetic manipulation. The term "phagemid" was coined separately in 1984 by Soviet scientists Melnikov et al. for phage-plasmid hybrids (e.g., λgt::pMB9, λNM::), which exhibited stability in lysogens and phage packaging in lytic cycles, yielding high transduction efficiencies (100- to 1,000-fold over standard particles). However, these -phagemids differ from the standard filamentous phagemids used in ssDNA generation and display, focusing instead on transduction.

Key Developments

Phage display technology was pioneered in 1985 by George P. Smith using full filamentous phage vectors (fd-tet derivatives with fusions to gene III), demonstrating display on virion surfaces for affinity selection. Phagemids were integrated into this system in the late 1980s and early 1990s to enable monovalent display, larger inserts, and higher library diversity. A key milestone was the 1990 work by McCafferty, Griffiths, , and colleagues, who used phagemid vectors to display functional fragments (Fd and scFv) on phage, facilitating selection of binders without and generating libraries >10^9 variants. In the 1990s, phagemids became standard for single-stranded DNA production in , with protocols like microtiter plate-based template preparation supporting projects. Late developments also included phagemid-based shuttle vectors for mycobacteria, enabling DNA introduction into species like M. smegmatis. From the , optimizations improved phagemid utility in high-throughput applications, such as larger insert capacities and better packaging. Reece's 2004 study highlighted phagemids for in functional protein analysis. Qi et al.'s 2012 review detailed phagemid properties for coat protein expression and library construction. A 2018 advancement was the pScaf phagemid, a customizable pUC18/M13mp18-derived vector producing ssDNA scaffolds (1,512–10,080 bases) for .

Molecular Structure

Essential Components

Phagemids are hybrid cloning vectors that integrate core plasmid elements to support high-copy replication and selection in bacterial hosts like Escherichia coli, enabling their use in DNA manipulation and library construction. These essential components form the foundational backbone, distinguishing phagemids from purely phage-based systems by providing stable double-stranded DNA maintenance without requiring helper phages for basic propagation. The origin of is a critical element, most commonly the ColE1-derived pUC origin, which drives high-copy-number replication (typically 300–500 copies per cell) of the double-stranded phagemid DNA during bacterial growth. This origin functions through RNA-primed synthesis initiated by host machinery, ensuring efficient amplification and ease of isolation for downstream applications. In representative vectors like pBluescript II, the ColE1 ori spans approximately 668 base pairs and supports robust propagation in standard E. coli strains. Selectable markers, usually antibiotic resistance genes, allow for the identification and maintenance of transformed cells under selective pressure. The ampicillin resistance gene (bla), encoding β-lactamase, is widely used, conferring resistance by hydrolyzing the in the periplasmic space and enabling selection on ampicillin-containing media. Alternatives like the kanamycin resistance gene (kan) may be employed in some constructs for broader compatibility, but ampicillin resistance remains standard due to its compatibility with high-copy plasmids. A (MCS), also known as a polylinker, provides a compact region with 15–25 unique restriction endonuclease sites for precise insertion of foreign DNA fragments. This facilitates , gene fusion, or the creation of diverse by allowing directional cloning without disrupting vector integrity. In pBluescript II, the MCS is embedded within the lacZα gene for blue-white screening, containing sites such as , , and NotI to accommodate various inserts up to several kilobases. Promoter elements are included to drive transcription of cloned genes or reporter sequences when expression is required, often inducible for controlled regulation. The lac promoter, responsive to (IPTG) in lacI^q strains, supports moderate-level expression suitable for toxic proteins or screening assays. Vectors like pBluescript II also feature T7 and T3 promoters flanking the MCS, enabling high-yield RNA transcription using phage polymerases, though these are optional for basic functions.

Replication and Packaging Elements

Phagemids incorporate phage-derived origins of replication, such as the f1 or M13 origin, which are essential for initiating the synthesis of single-stranded DNA (ssDNA). These origins contain specific sequences, including an ssDNA start site and termination site, that direct the host cell's RNA polymerase to produce RNA primers for ssDNA replication, distinguishing phagemid behavior from standard plasmid replication. The signal, located within the (IGR) of the f1 or M13 , serves as a recognition site for phage replication proteins to encapsidate the ssDNA into virions. This region, typically spanning about 400-500 base pairs, includes promoter elements and the packaging initiation site that ensure selective incorporation of phagemid DNA over other cellular nucleic acids during virion assembly. Some phagemid designs, particularly those for , include a cloning site for fusion of inserts to the III (gIII) encoding the minor protein pIII, enabling surface display of peptides or proteins. Other designs optionally include a partial or modified encoding a protein, such as gene VIII (gVIII) encoding the major protein pVIII, to provide a limited supply of structural proteins and thereby reduce dependence on helper phages for efficient packaging. This feature allows for hybrid virions where phagemid-encoded proteins compete with wild-type proteins from the helper, improving yield and purity in certain applications. Due to the fixed dimensions of the filamentous phage capsid, phagemid inserts are constrained to less than 10 kb in total length to ensure proper packaging and virion formation. Exceeding this limit disrupts capsid assembly, limiting the size of cloned DNA fragments.

Mechanism of Action

Double-Stranded Replication

Phagemids maintain their double-stranded form through a plasmid-like replication process in bacterial hosts such as Escherichia coli, utilizing a ColE1-derived origin of replication that operates independently of phage elements. This replication mode ensures the phagemid exists as a stable, extrachromosomal circular dsDNA molecule, relying solely on host cellular machinery for propagation. Initiation of replication begins at the origin, where host transcribes an II pre-primer from a promoter upstream of the origin. The RNA II molecule hybridizes to the template near the origin, forming a persistent hybrid structure, after which host RNase H cleaves the RNA to generate a primer with a free 3'-OH end. then extends this primer to initiate leading-strand synthesis in a unidirectional manner, with III taking over for processive elongation and lagging-strand synthesis via . This theta-structured replication produces daughter circular dsDNA molecules without requiring phage involvement. The replication process features relaxed control due to the regulatory interplay between RNA II and its antisense counterpart, RNA I, which inhibits primer maturation; in many phagemid vectors, mutations in the RNA II/RNA I interaction region reduce this inhibition, resulting in a high copy number of 50-300 phagemid molecules per host cell. This elevated copy number enhances yield and stability during . Phagemids are selected and propagated via integrated antibiotic resistance markers, such as the bla gene conferring resistance, which ensures only transformed cells survive under selective conditions. In the presence of helper phage, phage-encoded proteins can induce a switch to single-stranded DNA production, but double-stranded replication persists autonomously to maintain the plasmid population.

Single-Stranded DNA Production

Phagemids, which maintain as double-stranded plasmids in Escherichia coli, require superinfection with a helper phage, such as M13KO7, to initiate single-stranded DNA (ssDNA) production. The helper phage supplies essential viral proteins absent in the phagemid, including the minor coat proteins encoded by genes III, VI, VII, and IX, which are critical for virion assembly. These proteins enable the packaging of phagemid-derived ssDNA into phage-like particles, distinguishing this process from the phagemid's constitutive double-stranded replication mode. Upon , the f1 (ori) on the phagemid directs rolling-circle replication to generate the positive-sense (+) ssDNA strand. This mechanism involves nicking of the dsDNA at the f1 ori by the phage-encoded gene II protein, followed by displacement synthesis and circularization of the displaced strand, producing multiple copies of the phagemid genome as ssDNA. The resulting ssDNA is coated with gene V protein to stabilize it, preparing it for .90015-6) The ssDNA is then packaged into filamentous virions resembling M13 phage particles, with the major coat protein (gene VIII) forming the cylindrical sheath and minor coat proteins anchoring the ends. These virions are secreted from the host cell through a type II secretion system without causing , allowing continuous production over extended culture periods. Yields can reach up to 10^{12} particles per milliliter of culture under optimized conditions.

Applications

Single-Stranded DNA Generation

Phagemids played a pivotal role in generating single-stranded DNA (ssDNA) templates essential for during the pre-next-generation sequencing (NGS) era. Developed in the early 1980s as hybrid vectors combining and filamentous phage elements, phagemids enabled efficient production of ssDNA from Escherichia coli cultures, serving as optimal substrates for the dideoxy chain-termination method. This approach required clean ssDNA to avoid secondary structures that could complicate primer annealing and chain extension, making phagemid-derived templates a standard in labs for sequencing cloned inserts up to several kilobases. In , phagemid-produced ssDNA facilitates precise genetic modifications through hybridization. A mutagenic anneals to the ssDNA template, and extends the primer to create a mutated double-stranded , which is then transformed into for replication and selection. This method, refined by the Kunkel protocol using uracil-substituted templates to enrich for , achieves mutation efficiencies exceeding 50% and has been widely adopted for altering protein-coding sequences or regulatory elements. Phagemids' compatibility with such protocols stems from their ability to yield high-purity ssDNA suitable for manipulations. Compared to traditional M13 vectors, phagemids provide advantages in ssDNA production, including higher overall yields—often reaching 1–10 mg per liter of culture—and simpler handling due to their backbone, which supports high-copy-number replication of double-stranded DNA for initial and propagation. While M13 vectors require steps to transfer inserts, phagemids integrate these features, reducing labor and contamination risks during ssDNA via helper phage infection. Optimized protocols further enhance phagemid ssDNA purity and quantity by minimizing helper phage contamination. The rise of NGS technologies, which process double-stranded DNA directly, has led to a marked decline in phagemid use for routine sequencing and mutagenesis since the early 2000s, as these methods no longer necessitate ssDNA preparation. Nonetheless, phagemids retain niche utility in , particularly for synthesizing custom-length ssDNA scaffolds in applications, where precise sequence control enables folding into complex nanostructures for and biomaterial design. For instance, engineered phagemids like pScaf produce kilobase-scale ssDNA with minimal fixed regions, supporting scalable scaffold production for advanced assemblies. The mechanism of ssDNA generation in phagemids involves helper phage supplying packaging proteins that preferentially encapsidate the phagemid's origin-derived ssDNA, allowing its isolation from supernatants.

Phage Display and Protein Engineering

Phagemids enable the display of peptides or proteins on the surface of filamentous bacteriophages by fusing genetic inserts encoding these molecules to genes for phage coat proteins, such as the minor coat protein gIII (pIII) or the major coat protein gVIII (pVIII). This fusion is achieved through the insert into a phagemid vector that carries the relevant coat protein , allowing the expressed to be incorporated into the phage particle during , which is facilitated by a helper phage providing the necessary structural genes. The use of gIII fusions typically supports monovalent display for higher-affinity selections, while gVIII fusions allow multivalent display suitable for lower-affinity binders. In protein engineering, phagemid-based libraries are constructed by introducing diverse genetic variants into the phagemid vector, achieving library sizes of 10^9 to 10^11 unique variants, which are essential for affinity-based selections such as antibody discovery. These libraries link each displayed protein or peptide directly to its encoding DNA within the phagemid, enabling the recovery and propagation of selected variants through bacterial transformation and helper phage infection, with single-stranded DNA production playing a key role in maintaining library diversity during propagation. Such large-scale diversity facilitates the screening of vast combinatorial spaces, prioritizing high-affinity binders from immune or synthetic repertoires. The biopanning process, central to phagemid , involves iterative rounds of selection to enrich for desired binding specificities. In each round, the is incubated with an immobilized target antigen to allow binding, followed by rigorous washing to remove non-specific or low-affinity phages, of bound phages using competitive agents or shifts, and subsequent amplification in bacterial hosts to regenerate the enriched for the next . Typically, 3-5 rounds of biopanning progressively increase the proportion of high-affinity clones, with output titers monitored to assess enrichment. A prominent example of phagemid applications in therapeutic development is the discovery of (Humira®), the first fully human approved for clinical use, which was identified and optimized from libraries targeting tumor necrosis factor-alpha. This process involved constructing human antibody Fab libraries in phagemids, followed by biopanning to select high-affinity variants, demonstrating the technology's efficacy in generating biologics with sub-nanomolar affinities for treatment.

Other Applications

Beyond ssDNA production and phage display, phagemids are used in targeted systems. These involve engineering phagemid particles to deliver genetic payloads, such as CRISPR-Cas components, to specific cells for therapeutic purposes. For example, phagemid-based vectors have been developed for receptor-targeted transduction in mammalian cells and for creating antibacterial particles that produce toxins upon infection. Recent advances include non-replicative phagemid particles for CRISPR-Cas9 delivery to target in and miniaturized phagemids for ligand-displayed delivery, with applications in antimicrobial therapy and as of 2024.

Advantages and Limitations

Advantages

Phagemids offer dual replication modes, allowing them to function as plasmids for double-stranded DNA (dsDNA) propagation in Escherichia coli while also enabling single-stranded DNA (ssDNA) production and packaging into phage particles upon superinfection with a helper phage. This versatility eliminates the need for separate vectors to achieve both dsDNA cloning and ssDNA generation, streamlining workflows in molecular biology applications. Compared to full phage vectors, phagemids exhibit higher transformation efficiency into E. coli due to their smaller size, typically around 3-5 kb versus 8-10 kb for phages, which facilitates the introduction of larger and more diverse genetic libraries. For instance, phagemid-based libraries can achieve transformation efficiencies yielding up to 6.7 × 10⁹ members, significantly surpassing the 5 × 10⁸ members typical of phage vectors. The use of helper phage systems in phagemid propagation makes them cost-effective for constructing and screening large libraries, as the helper provides essential packaging proteins in trans, allowing efficient production of phagemid particles without the full genomic burden of a phage. This approach supports library sizes exceeding 10¹⁰ variants, reducing the expense and complexity associated with amplifying diverse populations for techniques like . Phagemids demonstrate strong compatibility with E. coli hosts, enabling straightforward genetic manipulation, high-yield plasmid amplification, and scalable phage production under standard laboratory conditions. This ease of handling in a well-characterized bacterial system enhances their utility across various biotechnological processes.

Limitations

Phagemids rely on co-infection with a helper phage to supply the necessary structural proteins for packaging into virions, which can result in incomplete infection and low packaging efficiency, often yielding only 1-10% of particles displaying the . This dependency also leads to variable ratios of helper phage to phagemid particles in preparations, potentially reducing selection efficiency and complicating high-throughput processes. Furthermore, the presence of helper phage introduces risks of genomic recombination between the phagemid and helper phage, generating wild-type phages that can outcompete phagemid particles and contaminate selections. The packaging of phagemid DNA into filamentous phage s imposes strict size constraints, typically limiting inserts to less than 10 kb due to the structural capacity of the , which accommodates genomes ranging from 4 to 12 kbp in total length. Exceeding this limit can impair virion assembly, infectivity, and , restricting phagemids to smaller constructs compared to some plasmid-based systems. In library construction for applications like , fusion of peptides or proteins to coat proteins can introduce biases, as toxic fusions may reduce rates or cause cell , leading to underrepresentation of those variants in the library. This skewing effect is exacerbated by expression variability in E. coli hosts, favoring non-toxic clones and diminishing library diversity. While phagemids remain valuable for display technologies, their use for single-stranded DNA production has become less common with the advent of next-generation sequencing and alternatives like enzymatic methods or PCR-based approaches; emerging systems such as yeast display offer advantages in handling larger proteins and eukaryotic folding without these prokaryotic biases.

References

  1. https://doi.org/10.1016/j.jmb.2012.01.038
  2. https://pmc.ncbi.nlm.nih.gov/articles/PMC11115567/
  3. https://pmc.ncbi.nlm.nih.gov/articles/PMC11680138/
  4. https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/phagemid
  5. https://pmc.ncbi.nlm.nih.gov/articles/PMC3044727/
  6. https://pmc.ncbi.nlm.nih.gov/articles/PMC101254/
  7. https://scholarworks.boisestate.edu/cgi/viewcontent.cgi?article=2250&context=icur
  8. https://www.sciencedirect.com/topics/medicine-and-dentistry/phagemid
  9. https://pubmed.ncbi.nlm.nih.gov/6300771/
  10. https://pubmed.ncbi.nlm.nih.gov/6234200/
  11. https://doi.org/10.1016/0378-1119(84)90084-2
  12. https://www.science.org/doi/10.1126/science.4001944
  13. https://www.nature.com/articles/347292a0
  14. https://pmc.ncbi.nlm.nih.gov/articles/PMC6784186/
  15. https://pmc.ncbi.nlm.nih.gov/articles/PMC328733/
  16. https://www.nature.com/articles/327532a0
  17. https://pubmed.ncbi.nlm.nih.gov/22310045/
  18. https://academic.oup.com/synbio/article/3/1/ysy015/5068535
  19. https://pubmed.ncbi.nlm.nih.gov/30984875/
  20. https://www.agilent.com/cs/library/usermanuals/public/212205.pdf
  21. https://pmc.ncbi.nlm.nih.gov/articles/PMC6461039/
  22. https://www.sciencedirect.com/topics/immunology-and-microbiology/phagemid
  23. https://pmc.ncbi.nlm.nih.gov/articles/PMC10334589/
  24. https://pmc.ncbi.nlm.nih.gov/articles/PMC55471/
  25. https://www.nature.com/articles/s41598-023-45087-2
  26. https://pmc.ncbi.nlm.nih.gov/articles/PMC9982796/
  27. https://pmc.ncbi.nlm.nih.gov/articles/PMC4251578/
  28. https://journals.asm.org/doi/10.1128/microbiolspec.plas-0029-2014
  29. https://www.cell.com/fulltext/0092-8674%2887%2990597-6
  30. https://pmc.ncbi.nlm.nih.gov/articles/PMC2846101/
  31. https://blog.addgene.org/plasmid-101-origin-of-replication
  32. https://pmc.ncbi.nlm.nih.gov/articles/PMC6024766/
  33. https://www.tandfonline.com/doi/full/10.2144/000112294
  34. https://link.springer.com/protocol/10.1385/1-59259-409-3:103
  35. https://pubmed.ncbi.nlm.nih.gov/2698664/
  36. https://conductscience.com/generation-of-single-stranded-dna-with-phagemid-vectors/
  37. https://pmc.ncbi.nlm.nih.gov/articles/PMC2692612/
  38. https://www.nature.com/articles/s41598-019-42665-1
  39. https://academic.oup.com/nar/article/30/5/e18/1016540
  40. https://www.pnas.org/doi/pdf/10.1073/pnas.88.18.7978
  41. https://www.sciencedirect.com/science/article/abs/pii/S0022283602005612
  42. https://pmc.ncbi.nlm.nih.gov/articles/PMC9369378/
  43. https://www.cellorigins.com/biopanning
  44. https://pmc.ncbi.nlm.nih.gov/articles/PMC5058633/
  45. https://www.frontiersin.org/journals/immunology/articles/10.3389/fimmu.2020.01986/full
  46. https://www.sciencedirect.com/science/article/pii/S1472979212001333
  47. https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0303555
  48. https://www.nature.com/articles/s42003-024-06754-w
  49. https://www.biorxiv.org/content/10.1101/2024.10.30.621196v1
  50. https://doi.org/10.1016/S0022-2836(02)00561-2
  51. https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0075212
  52. https://pmc.ncbi.nlm.nih.gov/articles/PMC1693883/
  53. https://academic.oup.com/femsre/article/39/4/465/2467559
  54. https://www.sciencedirect.com/science/article/abs/pii/S0378111906007700
  55. https://pmc.ncbi.nlm.nih.gov/articles/PMC2640155/
  56. https://academic.oup.com/femsre/article/46/2/fuab052/6407522
  57. https://journals.asm.org/doi/10.1128/microbiolspec.aid-0028-2014
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