HORDS Method for Obligate Lytic Phage Genome Modification
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Solution Overview
Problem
Current methods are inadequate for genetically modifying obligately lytic bacteriophages, as they cannot utilize conventional selection markers due to their obligate lytic lifestyle, requiring specific knowledge of phage and host cell genes involved in replication, and are not broadly applicable across bacterial species.
Innovation Solution
The HOst Range Determinant Selection (HORDS) method involves using a vector with a phage-targeting region containing a host range determinant from a marker phage, flanked by sequences homologous to the target phage genome, allowing for genetic modification and selection of modified phages without relying on host cell characteristics.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional selection markers are used for genetic modification, then selection efficiency is improved, but applicability to obligately lytic phages is worsened
Solution Approach 1:
The patent introduces a host range determinant as an intermediary selectable marker that bridges the gap between conventional selection methods and obligately lytic phage modification. This determinant allows selection of recombinant phages without requiring the phage to establish lysogeny or rely on host cell survival, thus enabling genetic modification of obligately lytic phages while maintaining efficient selection
Solution Approach 2:
The patent changes the selection parameter from host cell-based markers (which require lysogeny) to phage-based host range determinants. This parameter change allows selection to occur at the phage level rather than the host cell level, making the method applicable to obligately lytic phages that cannot integrate into host genomes
2Ease of operation
If standard molecular genetic techniques are used, then ease of manipulation is improved, but requirement for specific knowledge of phage and host genes is worsened
Solution Approach 1:
The patent creates a universal method for phage genetic modification that works across different phage types and bacterial species. The host range determinant selection system serves multiple functions: it enables selection of recombinants, works with obligately lytic phages, and does not require species-specific optimization, thereby simplifying the overall process while reducing the need for extensive specific knowledge
3Ease of manufacture
If temperate phage are used for genetic modification, then ease of genetic modification is improved, but ability to replicate lytically is worsened
Solution Approach 1:
The patent inverts the conventional approach by using host range determinants (typically associated with lytic phage infectivity) as selectable markers instead of using lysogeny-based selection. This inversion allows the modification process to work with obligately lytic phages while maintaining their lytic replication capability, effectively solving the contradiction by reversing the selection mechanism
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This method enables efficient genetic modification of lytic phages by selecting for host range determinants, allowing for the incorporation of exogenous DNA, mutations, or deletions, and is applicable to any bacterial species, rendering it broadly useful for modifying obligately lytic phages.
Implementation Method 1
a phage-targeting region which comprises a host range determinant of a marker phage, different from the host range determinant of the target phage, wherein the phage-targeting region is flanked by first and second flanking sequences homologous to the first and second target sequences of the target phage genome
Data Source
AI summary
A method for modifying the genome of a lytic target phage, uses of the method and products thereof are described. Compositions comprising such phage are also described. The compositions may be formulated as a medicament, which are useful for human treatment and may treat various conditions, including bacterial infections.