Custom Herpes Simplex Virus Vector Production Service
Overview of Herpes Simplex Virus Vector
There are two subtypes of the herpes simplex virus (HSV): HSV-1 and HSV-2. It belongs to the family Herpesviridae. The HSV genome is made up of covalently linked long fragments (UL) and short fragments (US) and is a linear double-stranded DNA of about 152 kb. A single sequence and inverted repetition sequences are present in every fragment. Figure 1 depicts the structure of the virion particle and the organization of the HSV-1 genome. The genome will cycle after the virus has invaded the host cell. Three unique characteristics spark interest in HSV-1-based vectors.
- The first is the viral particle's enormous size, which enables the effective delivery of up to 150 kbp of DNA and lowers the danger of mutagenesis by preventing the vector genome from integrating with host DNA.
- The complexity of the virus genome, which contains about 40 genes that are not necessary for virus replication in cell culture and may be removed without impairing virus production, is the second distinguishing trait.
- The third is the remarkable adaptations of HSV-1 to the nervous system, which include the ability to regulate neurovirulence, spread anterogradely and retrogradely between neurons, and establish latency in neurons, a nontoxic state that permits transgenic transcription under the control of suitable promoters.
Fig. 1 Organization of the HSV-1 genome and structure of the virion particle.1
Most HSV vectors are developed from strains of HSV-1. HSV-1 has been used to create three different kinds of vectors: Amplicon vectors, Defective Recombinant Vectors, and Amplicon Vectors.
Attenuated Vectors are replication-competent viruses that have gene alterations that are critical for virulence in the injected hosts but are not necessary for virus multiplication in cultured cells. These genes typically participate in a variety of interactions with cellular proteins that enhance the virus' capacity to proliferate within cells or elude host defenses.
Defective Recombinant Vectors are disabled, replication-incompetent, non-pathogenic vectors that lack one or more essential IE genes. These vectors cannot spread within the host but retain many advantageous features of the wild-type virus, particularly the ability to express transgenes after establishing latent infections in central and peripheral neurons following local administration.
Amplicon Vectors are HSV-1 particles that are identical to wild-type HSV-1 but that instead carry the amplicon plasmid, a concatemeric version of a DNA plasmid. Amplicons are special in that they may supply whole genomic regions, including substantial regulatory elements and introns, and then transform them into synthetic human chromosomes.
Production
HSVs produced by Creative Biolabs are either devoid of assistance particles or have minimal to no helper particle contamination. One method is the cotransfection of amplicon plasmids and BACs (BAC-HSV), which deliver the entire complement of transacting HSV-1 activities. When compared to the HSV-1 genome of the wild type, the HSV-1 helper genome given by the BAC is significantly bigger and absent of viral packaging signals (pac). Helper-free amplicons are created as a result of these helper genomes' failure to wrap around freshly formed HSV-1 particles. Another relies on Cre/loxP1-based site-specific recombination to eliminate the helper virus packing signals in the cells producing the amplicons. Recombinant HSV-1 is used in this system as a helper since it has an ectopic pac signal and two parallel loxP sites on either side of it. Since the pac signal was removed, this helper virus' genome has become Cre-sensitive and will not package in cells that express Cre.
Purification
To further purify and concentrate viral particles for ultra-purified HSVs, sucrose density gradient centrifugation is commonly used. We can also offer additional purifying techniques. Please contact us if your purification needs are higher so we can offer you a tailored purification solution.
Quality Control
Our quality control service for HSV vectors includes Titer tests, Sterility tests, Mycoplasma tests, and Fluorescent transduction tests if the HSV vectors express a fluorescent protein. For ultra-purified HSV, we provide Endotoxin tests additionally. One of the world's top innovators in vaccination technology development, Creative Biolabs offers the best and most complete services for HSV vector production. Please contact us for more information.
Reference
- Taylor, Travis J et al. "Herpes simplex virus." Frontiers in bioscience : a journal and virtual library vol. 7 d752-64. 1 Mar. 2002, doi:10.2741/taylor.
