Question

2) (10 point question) FGF receptors are known to have more than one isoform due to alternative splicing. See Figure 2 beld Acid FGFR2 Illb Box II encoded protein Kinasel Kinase2 Illa CS TM Fgfr2 allele Fgfr2 alete -5-6- 6 1 0 Acid Box II FGFR2 Ilic encoded protein TM Kinasel Kinase2 Figure 2 Alternative splicing, transcription and isoforms of FGFR2 The N-terminal half of the third Ig loop is encoded by exon 7, while the C-terminal half of the third Ig loop is encoded by either exon 8 or 9, which generate the alternative splicing isoforms, IIIb and IIIc. Exon 10 encodes the transmembrane domain. I, II and III represent the three Ig loops located in the extracellular domain; TM, transmembrane domain; S, secretory peptide. You're recruited by a research lab to make a conditional allele of Fgfr2 in mice. Their aim is to research the role of different FGFR protein variants in short-stature syndromes. This allele will serve two purposes: 1) They need to be able to switch-on FGFR2 in the chondrocytes only. 2) They only want to express the FGFR2 IIIb isoform. A) (4 points) Draw an rough diagram of an HDR template, including the appropriate parts of the FGFR2 gene, that you will use with a knock-in strategy to modify the Fgfr2 gene. Label all the parts you need.

Answer 1

To answer this question, you have to understand the concept of conditional allele and how it is created. Cre-lox or FLP-FRT recombinase system are the keys of gene site-specific gene editing in eukaryotic systems.

Bore recombinase and its close relative FLP recombinase are both used experimentally to delete genes in eukaryotic organisms. Cre is an enzyme encoded by phage P1 , which functions to circularize the linear phage genome during infection. The recombination sites on the DNA, where are acts, are called lox sites,

The diagram of the FGFR2 allele is hereby given

EHH484ah101 LOX P Conditional alleles are alleles that are modified according to our own wish. Inserting ton P sites before and after a exon a will flose exon a on treating the mice with cre recombinase. Until treatment, the nice mouse is capable of synthesizing both FG FR2 II b and ille. However, when the recombinase is present, only FG FR 2 IIIb will be synthesized. The

The first question was this allele has to be chondrocyte specific. Targeted Cre recombinase should be applicable in this case. Every cell (including chondrocytes) have specific markers or transcription markers, which can be tagged with the Cre recombinase system.