03 - Chick Check: The Genetics Behind Blue Chicken Eggs
we want to copy. Finally, the Taq polymerase uses the nucleotide building blocks to synthesize new DNA strands. This cycle – denature, anneal, extend – forms the core of PCR and allows us to quickly create billions of copies of our target sequence. To detect the presence or absence of the EAV-HP viral insertion, we use a clever 3-primer PCR approach (Figure 5): • Forward Primer Non-Blue Egg: Binds upstream of the viral insertion site. • Reverse Primer Non-Blue Egg: Binds downstream of the insertion site. • Internal EAV-HP Primer: Binds within the EAV-HP viral insertion sequence that creates the “blue egg gene.” This creates different band patterns depending on the chicken’s genotype. • Homozygous (two copies) Non-Blue Egg Chicken: One band (~283 bp) from Forward and Reverse Primers for Non-Blue Egg. • Homozygous (two copies) Blue Egg Chicken: One band (~637 bp) from Forward Non-Blue Egg and Reverse EAV-HP Primers. • Heterozygous (One copy of each gene) Blue Egg Chicken: Two bands (both ~283 bp and ~637 bp) from all three primers.
Forward Primer
A
Eggshell gene
Reverse Primer
283bp
EAV-HP viral insertion
Internal EAV-HP Primer
Forward Primer
B
Eggshell gene
Reverse Primer
637bp
Lane 1
Lane 2
Lane 3
637 bp
C
283 bp
Figure 5: The 3-Primer PCR system. (A) DNA without viral insertion showing non-blue primer binding. (B) DNA with viral insertion showing internal primer binding. (C) A simulated electrophoresis gel showing homozygous non-blue eggs (Lane 1), homozy- gous blue eggs (Lane 2), and heterozygous blue eggs (Lane 3). Eggs with the insert produce a 637 bp band & eggs without the insert produce a 283 bp band.
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