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Lentivirus

 

Custom engineered Lentivirus vectors for gene overexpression and knockdown 

 Lentiviruses are the most versatile method for gene delivery. Lentiviruses infect the broadest panel of cells of dividing and non-dividing cells, including primary cells, immortalized cell lines, hematopoietic stem cells. Thanks to a large insert site for an expression cassette, lentivirus offers flexible vector design, including insertion of several genes with a total capacity up to 5 kb. Lentivirus become a tool of choice
whenever a fast, efficient and cost-effective generation of stable homogeneous cell models is required, either with a strong gene expression or when knockdown is needed. In gene therapy Lentivirus are widely used for CAR-T cell therapy applications.

 SIRION Biotech’s custom lentivirus service provides constitutive and inducible vectors for overexpression and gene knockdown. Our multicistronic vector design results in highly homogeneous, stable cell models avoiding time-consuming clonal selection posttransduction. With the all-in-one inducible vector system SIRION enables working with difficult, cytotoxic genes and shRNA strategies. We offer vectors in less than 4 weeks.

Custom Lentivirus at a glance:

Every project starts with personal project consultancy with our specialists

  • Application field from in vitro to in vivo
  • Stable gene expression due to integration of lentivirus into host genome
  • Reliable gene delivery into dividing and non-dividing cells
  • Large packaging capacity
  • 3rd generation SIN vectors for higher safety
  • Highly homogeneous stable cell pools through optimized vector design
  • Mono- and bicistronic vector design
  • Capsid retargeting options
  • Broad selection of promotors, selection markers and fluorescent tags
  • Reliable gene delivery into dividing and non-dividing cells
  • Inducible LV technology enabling characterization of toxic gene modulations based on TetOn and TetOff systems
  • Lentiviral particles titer variety from small to extra large
  • Thorough quality control and verification of physical and functional titers

3’SIN Technology Lentivirus vectors

To ensure the safety of Lentivirus vectors and prevent virus generation, 3rd-generation lentivectors exhibit the viral genome divided into separate plasmids. SIRION Biotech uses 3’SIN Technology (self-inactivative) of 3rd generation Lentivirus vector engineering. In 3’SIN Technology 400 bp deletion in the U3 region of the 3‘UTR inactivates promoter activity. During integration of the viral DNA into the host genome the 3‘SIN LTR is transferred to the 5‘end resulting in a provirus lacking U3 promoter activity at both ends. 3’SIN Technology enables controlled gene expression from internal promoter(s) which is a mandatory prerequisite for all inducible, cell type-specific or pathway reporter expression systems.

  • No overlapping sequences between the co-transfected vectors to eliminate any homologous recombination
  • VSV-G protein pseudotyping for broad host tropism
  • Controlled gene expression from internal promoter(s)

In 3’SIN 3 packaging plasmids are used to express all protein components necessary for LV packaging and generation of infectious viral particles plus a plasmid carrying GOI. Lentivirus vectors are pseudotyped with VSV-G protein for mediation of fusion of the viral envelope with the host cell.

Constitutive Lentivirus

Depending on the project needs, it can be necessary to design a construct with strong constitutive gene overexpression or to induce expression at a particular time point. In this case it is important to design the construct which fits your needs the best. SIRION Biotech offers a broad portfolio of constitutive and inducible promoters for Lentivirus vectors.

  • Stable cell model generation to analyze long-term effects of gene overexpression and silencing in vitro
  • For dividing and non-dividing cells
  • Variety of strong and ubiquitous promoters (CMV, Ubi, EF1-α, etc.)
  • Custom promoters
  • IRES or 2A sequences for bicistronic vectors
  • Knockdown efficiency of at least 80% on mRNA level when combining with our RNAiONE service

Service Details

  • Cloning of customer cDNA/shRNA in SIRION’s lentiviral expression vector
  • Verification of cloning success by DNA-Sequencing
  • Lentivirus production in HEK293 cells
  • Purification, concentration and QC
  • Quality control: physical particles determination (GC titer),  functional titers (TU titer, IU titer).

Inducible Lentivirus

SIRION Biotech’s inducible expression platform is based on the latest 3G TET technology. This guarantees a high sensitivity without leakiness. SIRION offers inducible lentiviral ONE-vector and TWO-vector systems.

Fields of Application

  • Establish stable inducible cell models
  • Circumvent cell adaption to stable genetic modification
  • Enable characterization of toxic gene modulations

SIRION’s benefits

  • TET-inducible All-in-ONE vector backbone
  • TET-inducible TWO-vector technology for large constructs
  • Highly sensitive without leakiness
  • Knockdown efficiency of at least 80% on mRNA level when used in combination with RNAiONE
  • Custom lentivirus in less than 4 weeks

Service Details

  • Cloning of your cDNA/shRNA into an inducible lentiviral expression vector
  • Verification of cloning success by DNA-Sequencing
  • Lentivirus production in HEK293 cells at different scales
  • Purification, concentration and QC
  • Quality control: physical particles determination (GC titer),  functional titers (TU titer, IU titer).

Contact us – We are looking forward to discuss your project.