MMLV Retrovirus Packaging
Recombinant Moloney murine leukemia virus (MMLV) retroviral vectors serve as efficient viral vector tools for introducing genes permanently into a wide variety of dividing cells. This retroviral system came into prominence as a gene delivery method for generating iPS cells.
VectorBuilder has developed a series of proprietary technologies and reagents that have greatly improved recombinant MMLV retrovirus production protocols in terms of titer, purity, viability and consistency, especially for the MMLV vector systems used in our vector cloning services. As a result, we have a growing base of highly satisfied customers who come back to us time after time for their MMLV vector cloning and MMLV packaging needs.
Types of MMLV retrovirus offered
- VSV-G pseudotyped wildtype MMLV retrovirus
- VSV-G pseudotyped self-inactivating MMLV retrovirus
Service Details
Price and turnaround Price Match
| Scale | Application | Titer | Volume | Price (USD) | Turnaround |
|---|---|---|---|---|---|
| Pilot | Cell culture | >107 TU/ml | 250 ul (10x25 ul) | $449 |
6-12 days
|
| Medium | 1 ml (10x100 ul) | $649 | |||
| Large | >108 TU/ml | 1 ml (10x100 ul) | $1,099 | ||
| Ultra-purified large | Cell culture & in vivo | >108 TU/ml | 1 ml (10x100 ul) | $1,699 |
TU = Transduction units (aka infectious units)
Deliverables
| For non-ultra-purified scales | For ultra-purified scales |
|---|---|
| Your custom MMLV retrovirus | Your custom MMLV retrovirus |
|
Free: control virus
|
Add-on purchase (optional): ultra-purified control virus
|
| Free: Polybrene (5 mg/ml, 200 ul) | Free: Polybrene (5 mg/ml, 200 ul) |
Control virus
The control retrovirus vector is designed to match the biological application of the custom virus and to be used for testing retroviral transduction. For example, if the custom virus overexpresses a gene, then the control virus provided will be EGFP control retrovirus (MMLV retrovirus overexpressing EGFP). Detailed information for the various control viruses is shown below:
Shipping and storage
Our MMLV retrovirus is stored in HBSS buffer and is shipped on dry ice. Upon receiving, it should be stored at -80°C for long term (stable for at least 6 months), or -20°C for use within one week. The shelf life for MMLV retrovirus is approximately one year. Please avoid repeated freeze-thaw cycles of lentivirus, as this can result in a large titer drop.
Technical Information
MMLV retrovirus production and quality control (QC)
For our MMLV retrovirus manufacturing, the transfer vector carrying the gene of interest (GOI) is co-transfected into packaging cells with our proprietary envelope vector encoding VSV-G and our packaging vector encoding Gag and Pol. After a short incubation period, the supernatant is collected and centrifuged to remove cell debris and then filtered. Viral particles are subsequently concentrated with PEG. For ultra-purified MMLV retrovirus (in vivo grade), viral particles are further purified and concentrated by sucrose gradient ultracentrifugation. We use a qPCR-based approach to measure the functional titer of MMLV.
For each recombinant MMLV retrovirus produced by VectorBuilder, quality control includes titer measurement, sterility testing for bacteria and fungi, and mycoplasma detection. If the transfer vector encodes a fluorescent protein, we would transduce cells with virus and detect corresponding fluorescence. Additionally, for ultra-purified MMLV, we routinely sample virus quality using the endotoxin assay. To include the endotoxin results in your COA, an extra cost is required. Additional QC services can be provided upon request.
How to Order
If customer-supplied MMLV retroviral plasmids are needed, please send them to us following the Materials Submission Guidelines. Please strictly follow our guidelines to set up shipment to avoid any delay or damage of materials. All customer-supplied materials undergo mandatory QC by VectorBuilder which may incur a $100 surcharge for each item. Please note that production may not be initiated until customer-supplied materials pass QC.
Resources
FAQ
| Lentivirus | MMLV | Adenovirus | AAV | |
|---|---|---|---|---|
| Tropism | Broad | Broad | Ineffective for some cells | Depending on viral serotype |
| Can infect non-dividing cells? | Yes | No | Yes | Yes |
| Stable integration or transient | Stable integration | Stable integration | Transient, episomal | Transient, episomal |
| Maximum titer | High | Moderate | High | Very high |
| Promoter customization | Yes | No | Yes | Yes |
| Primary use | Cell culture and in vivo | Cell culture and in vivo | In vivo | In vivo |
| Immune response in vivo | Low | Low | High | Very low |
Our titer guarantee only applies to vectors for which the region being packaged into virus (from 5’ MoMuLV LTR to 3’ MoMuLV LTR) is below MMLV cargo limit (8.3 kb). For sizes above this, it may still be possible to package the vector into virus, but the titer may be reduced. Additionally, we are not able to guarantee titer for the following vectors:
- Vectors containing sequences that could adversely affect the packaging process such as toxic genes (e.g. proapoptotic genes), genes that compromise the integrity of packaging cells or virus (e.g. membrane proteins that cause cell aggregation), and sequences prone to rearrangements or secondary structures (e.g. repetitive or highly GC-rich sequences);
- Customer-supplied plasmids containing undisclosed sequences or atypical MMLV functional elements (e.g. LTR) that may introduce uncertainties to packaging efficiency.
Our estimated turnaround is the time from production initiation to completion. It does not include waiting time for any customer-supplied materials (e.g. template DNA or viral vectors), QC of such materials, and transit time for shipping final deliverables to the customer.