Stable Cell Line Generation

VectorBuilder can provide stable cell line production services with desired genetic modifications at unbeatable prices and rapid turnaround. VectorBuilder can design and generate customized vectors based on your experimental needs and the characteristics of your specific cell lines. Chemical transfection, electroporation or viral transduction are employed as appropriate to introduce the desired DNA changes into the target cells. Following standard drug selection, polyclonal cell populations or monoclonal cell lines are sub-cultured, verified and delivered to you ready for use. All cells are guaranteed with desired genotypes, contaminant-free and authenticated according to ATCC standards.

Types of stable cell lines offered
  • Overexpression stable cell lines
  • shRNA knockdown stable cell lines
  • Tet inducible gene expression stable cell lines
  • CRISPR stable cell lines (includes gene knockout, knockin and point mutation)

Service Details

Price and turnaround
Stable Cell Line Model Strategy Deliverable Price (USD) Turnaround
Gene Overexpression Lentivirus Based Pooled Cells From $3,199 9-15 weeks
Single Clone From $5,199 12-20 weeks
shRNA Gene Knockdown Lentivirus Based Pooled Cells From $5,699 8-13 weeks
Single Clone From $7,699 11-18 weeks
Tet Inducible Gene Expression Lentivirus-based Pooled Cells From $3,599 10-16 weeks
Single Clones From $5,899 13-21 weeks
Gene Knockout Regular Plasmid CRISPR/Cas9 Based Single Clone From $5,999 10-16 weeks
Lentiviral CRISPR/Cas9 Based Single Clone From $6,999 10-17 weeks
Gene Knockin Regular Plasmid CRISPR/Cas9 Based Single Clone From $10,099 14-19 weeks
Point Mutation Regular Plasmid CRISPR/Cas9 Based Single Clone From $9,199 13-19 weeks

Technical Information

Gene overexpression

VectorBuilder can generate stable cell lines overexpressing your gene(s) of interest (GOIs) to help you overcome variations associated with transient transfection. Overexpression stable cell lines are generated by lentiviral transduction followed by standard drug selection. The GOI expression level is validated by RT-qPCR. Additionally, a series of standard QC assays such as sterility tests and mycoplasma detection are performed for releasing the final cell line products. Other validation assays such as Western blot and immunocytochemistry can be performed upon request.

Gene Overexpression image
shRNA gene knockdown

VectorBuilder can generate shRNA knockdown stable cell lines for applications requiring long-term knockdown of your GOI(s). To ensure efficient knockdown of your GOI(s), the target cells are separately transduced with three lentiviruses carrying the top 3 shRNAs based on knockdown score. The cell line showing the best knockdown efficiency validated by RT-qPCR will be delivered. Additionally, a series of standard QC assays such as sterility tests and mycoplasma detection are performed for releasing the final cell line products.

shRNA gene knockdown image
Tet inducible gene expression

VectorBuilder can custom build Tet inducible gene expression stable cell lines with minimal background expression and high induction of your GOI(s). Additionally, we can generate Tet regulatory protein expression stable cell lines (e.g. rtTA, tTS/rtTA, Tet3G, etc.) which can then be transfected/transduced with plasmids/viruses carrying TRE driven GOI(s) for flexible experimental design and reliable Tet inducible gene expression. For Tet-On stable cell lines, induction of the GOI is validated by RT-qPCR. Additionally, a series of standard QC assays such as sterility tests and mycoplasma detection are performed for releasing the final cell line products.

Tet inducible gene expression image
Gene knockout

VectorBuilder can generate stable cell lines with permanent knockout of your GOI(s). Stable knockout of GOI(s) is achieved using a CRISPR-based approach, wherein a pair of gRNAs targeting the GOI(s) is introduced into the target cells along with Cas9. This leads to the generation of two cuts on the target gene. Attempts by cells to repair the broken ends of the two cut sites would typically lead to a large deletion spanning the two sites. The two cut sites are designed to flank a functionally important region of the target gene such that a deletion spanning them would most likely lead to the loss of gene function.

Depending on the cell type, either lentiviral or regular plasmid CRISPR vectors may be used for achieving target gene knockout. The final cell line is evaluated by Sanger sequencing of the targeted locus to confirm knockout. Additionally, a series of standard QC assays such as sterility tests and mycoplasma detection are performed for releasing the final cell line products.

Gene knockout image
Gene knockin

VectorBuilder can generate stable cells line with permanent knockin of your GOI(s) at desired genomic target sites. Stable knockin of GOI(s) is achieved by a CRISPR-based approach wherein a target site specific gRNA is introduced into the cells along with Cas9 and a donor vector which serves as the template for HDR-mediated gene knockin.

The final cell line is evaluated by Sanger sequencing of the targeted locus to confirm knockin. Additionally, a series of standard QC assays such as sterility tests and mycoplasma detection are performed for releasing the final cell line products.

Gene knockin image
Point mutation

VectorBuilder can generate stable cell lines harboring desired point mutations at genomic target sites of interest. Stable cell lines with point mutations are generated using a CRISPR-based approach wherein a target site specific gRNA is introduced into the cells along with Cas9 and a single-stranded donor oligonucleotide (ssODN) which serves as the template for HDR-mediated insertion of the point mutation.

The final cell line is evaluated by Sanger sequencing of the targeted locus to confirm the presence of the point mutation. Additionally, a series of standard QC assays such as sterility tests and mycoplasma detection are performed for releasing the final cell line products.

Point mutation image

How to Order

FAQ

Can VectorBuilder QC final cell lines by assays such as Western blot, immunocytochemistry and Southern
blot?

Stable cell lines generated by VectorBuilder undergo stringent QC procedures which typically include standard assays such as sterility tests and mycoplasma detection along with appropriate assays to validate the genotype of the final cell lines depending on the type of stable cell line. For gene overexpression, shRNA gene knockdown and Tet inducible stable cell lines RT-qPCR is performed for validating ectopic GOI expression, loss of GOI expression and induction of GOI expression in the final cell line, respectively. Whereas for CRISPR stable cell lines (gene knockout, knockin and point mutation), Sanger sequencing of the targeted region is performed to validate the presence of the desired genomic edit in the final cell line.

In addition to the standard QC assays mentioned above, VectorBuilder can perform Western blot, immunocytochemistry and Southern blot analyses on generated stable cell lines upon request. These assays are available as add-on services and can be offered after evaluating the feasibility of requested assays for the corresponding stable cell lines. However, VectorBuilder does not necessarily guarantee the results of the add-on validation assays, because some assays heavily rely on the availability of validated antibodies and suitable testing system (e.g. cell line, etc.).

How does VectorBuilder define turnaround time for stable cell lines?

For stable cell line generation services, VectorBuilder defines estimated turnaround as the time from production initiation to completion. Production completion is achieved once the final cell line passes QC and is ready to be shipped out. Turnaround time does not include waiting time for any customer-supplied materials (e.g. cell lines or template DNA for vector construction), QC of such materials, and transit time for shipping final deliverables to the customer.