To rescue native morphology of membranebound, forming VLPs. For this co-expression to be consistent, we show that single cell measurements of VLP assembly require a quantitative read-out for the presence of unlabeled Gag. Furthermore, in transient co-transfection the simultaneous uptake and expression of both plasmids is not guaranteed at the single cell level, which enforces the need for such a read-out. As a consequence, the co-transfection ratio of unlabeled to labeled Gag plasmid of 10 to 20 fold that has been previously proposed is only meaningful with such a quantitative read-out, given that the unlabeled to labeled Gag ratio can be subject to fluctuations over an order of magnitude. The ability to direct ES and induced pluripotent stem cell differentiation toward specific tissue fates in vitro provides an excellent opportunity to investigate the gene regulatory networks that operate during organ development. While ES and iPS cells hold promise for cell-based therapies, only in a handful of cases is molecular information detailed enough to guide directed differentiation to specific tissue types. The developing vertebrate ocular lens offers a potential system for such approaches, as considerable knowledge exists about the cascade of transcription factors, signaling molecules and cell-cell interactions necessary for head surface ectoderm to develop into a mature optically clear lens. This process is accompanied by the stepwise specification of the preplacodal region into an anterior sensory placode domain and then a pseudostratified ectodermal lens placode. Thereafter, progression through the lens pit and lens vesicle stages occurs, culminating in formation of the lens proper. From this stage on, the lens consists of anteriorly localized cells, termed the anterior epithelium of the lens, that terminally differentiate into posteriorly localized elongated fiber cells. Numerous studies demonstrate that lens differentiation involves the action of a conserved GRN that is initiated by a specific set of regulatory genes that includes Pax6 and Six3. Targeted mis-expression in Drosophila of mouse or fly Pax6 that encodes a conserved paired domain and homeodomain containing transcription factor results in multiple ectopic ommatidial structures on the antenna, wings and halteres. In addition, Pax6 mis-expression in Xenopus results in ectopic eye structures that include lens-like tissue termed “lentoids”, as well as retinal tissue. The formation of ectopic lentoids in the nasal periocular ectoderm is also noted in mice with conditional deletion of betacatenin, suggesting that canonical Wnt signaling normally represses lens fate. Thus, repression of canonical Wnt signaling in the surface ectoderm is critical for lens development, and Pax6 has been demonstrated to directly control expression of several Wnt Tofacitinib JAK inhibitor inhibitors in the presumptive lens ectoderm. Conversely, Pax6 haploinsufficiency in mice results in the Small eye and cataract phenotypes, and nullizygosity results in a failure of lens placode induction and anophthalmia. Similarly, PAX6 haploinsufficiency in humans results.