Primary PCR of antibody genes

Mouse heavy chain constant region primers 

Final assembly wKh primers contalninB restriction sites  

Set up the following 100 jil reactions for each PCR and negative control, i.e. one reaction for each of the nine family-specific Vh BACK primers (VhI to VhII Back primers, see TabU 1) and eight family-specific V BACK primers (V l to VrB Back, see Tabic 1), together with controls (no DNA) for each a total of 34 reactions. 

Add 1 pi cDNA mRNA hybrid (from Protocol 4). and overlay with 100 pi of mineral oil.  

Add 1 pi Deep Vents NA polymerase to each reaction beneath the mineral oil. using separate pipette tips for each addition. 

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PCR with primers for the two related genes indicated that both PAC a and PAC P were expressed in a number of phototaxis mutant strains, indicating that these are not primary photoreceptor mutants, but downstream mutants (unpublished data). None of the two proteins was detected in Astasia a nonphotosynthetic relative of Euj kna which lacks the PAB and consequently does not show photo taxis. Polyclonal antibodies were raised against these two proteins subunits and found to bind at the site of the PAB in the cell, indicating that these represent the photoreceptor. 

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