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 MMLV retrovirus

Service Details

Price and turnaround
Scale Application Titer Volume Price (USD) Turnaround
Pilot Cell culture >107 TU/ml 250 ul (10x25 ul) $399 8-16 days
Medium 1 ml (10x100 ul) $599
Large >108 TU/ml 1 ml (10x100 ul) $999
Ultra-purified large Cell culture & in vivo >108 TU/ml 1 ml (10x100 ul) $1,499

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

  • Pilot: >107 TU/ml, 100 ul
  • Medium: >107 TU/ml, 2x100 ul
  • Large: >107 TU/ml, 2x100 ul

  Add-on purchase (optional): ultra-purified control virus

  • Ultra-purified large: >108 TU/ml, 10x100 ul, at $500
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:

Vector system Control virus vector name Control virus vector ID
MMLV gene expression system pMMLV[Exp]-EGFP/Puro VB170316-1032yra
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 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 cushion centrifugation. We use a qPCR-based approach to measure the functional titer of MMLV.

MMLV-img

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.

Technical documents
User Instructions Material Safety Data Sheet (MSDS) Certificate of Analysis (COA)

How to Order

Order both vector construction and virus packaging
Order virus packaging or your own vector

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.

FAQ

What features does MMLV retrovirus have comparing to other viral vectors?
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 Very High 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
What is VectorBuilder's guarantee for virus titer and turnaround?

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 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.