Perhaps the scientists who wrote the attached paper would be able to help. They have the CDK4 null mouse. Making embryonic fibroblasts from 13.5 -14.5 dpc embryos is relatively easy but is a chore that few people like to do for free (I should know, I have done it many times and it is time consuming). You would need to go to their institute to prepare them (takes about 4-6 weeks including breeding time), or have the live mice sent to you so that you can breed them yourself and collect embryos at the right stage of development (can take 6-12 months), or ask the researchers with the knockout mice to make the fibroblasts for you and send you vials of frozen cells (if they have the experience). The latter is the easiest method from your work time point of view but you are depending on them to do it when they can spare the time, and you will need to offer them something in return.

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3973114/

Hello

You can follow this protocol The proposal by the Xianghong Zou

A targeted null mutation of the Cdk4 gene,Cdk4tm1Kiyo , was created in mouse embryonic stem cells, and mice with germline transmission of this mutation were bred in the recombinant C57BL/6 × 129/svj strain background, . MEFs were prepared from day-12.5 mouse embryos and cultured in the Dulbecco's modified minimum essential medium supplemented with 2 mM glutamine, 100 U/mL penicillin and streptomycin, and 10% fetal bovine serum (FBS; Life Technology), as described (Tsutsui et al. 1999). MEFs dispersed from each embryo using 0.25% trypsin solution containing 0.53 mM EDTA were cultured in a 100-mm culture dish (passage 1). Cells were then maintained using a 3T3 protocol (3 × 105 cells per 60-mm culture dish passaged every 3 d). The population doubling level during each passage was calculated according to the formula log(final cell number/3 × 105)/log2.

Good Luck 

Many thanks to both of you,

I anticipated creating the MEFs de novo would be quite a lot of work and was hoping there may be, by now, a way of purchasing some.

The alternative would be SiRNA CDK4 knockdown in MRC-5 cells to provide the null background.  Does anyone have any experience of this?

As a follow up to my earlier question:  Many gene knockouts including Cdk4 are now available in the in-vitro haploid immortal fibroblast cell  line HAP1.

These can be obtained from Horizon Discovery, Building 7100, Cambridge Research Park Cambridge CB25 9BL  UK