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#gene excision

2 public questions tagged with this topic.

Gene Tar getting

What would result from placing loxP sites flanking exon 2 of a gene in same orientation?

Placing loxP sites in same orientation flanking exon 2 and intronic regions preserves gene function until Cre acts. Cre recombinase synapses two directly repeated loxP sites, forming a loop and catalyzing strand exchange. Recombination excises the looped DNA as circular molecule containing exon 2 and one loxP site, leaving single loxP scar in chromosome. Loss of coding exon causes frameshift mediated nonsense or deletion of essential domain, achieving conditional knockout after tissue specific Cre activation without affecting other tissues developmentally.

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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Gene Tar getting

In conditional knockout using Cre, gene excision happens when:

Conditional knockout overcomes embryonic lethality by floxing essential exons with loxP sites placed in same orientation flanking critical region intronically, preserving normal splicing without Cre. Excision occurs only upon Cre expression which recombines directly repeated loxP sites, deleting intervening exon and creating frameshift or null allele. Tissue specific or tamoxifen inducible Cre drivers then restrict deletion temporally and spatially. Requirement of same direction loxP ensures deletion rather than inversion, producing irreversible loss of function in selected cells only in vivo efficiently.

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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