Mobilizing an insert to generate additional transformed lines.
(modified 6/18/99)

The potential for chromosomal position effects requires that we analyze several insertions before drawing conclusions about the behavior of a promoter construct. If one line can be obtained, it is possible to generate additional lines by performing a dysgenic cross with a flies containing the defective P-element called P[ry+q2-3](99B) - abbreviated q2,3 for this discussion. If the insert is on the X or 2nd chromosome, it is probably best to use the line X[yw]/X[yw]; Sp/Sm6a; Sbq2,3/Tm6Ubx. Note that the 3rd chromosome called Sbq2,3 is not a balancer chromosome so it will recombine with a wild-type chromosome. It is stable over TM6Ubx because Sb is homozygous lethal.  However, when doing the dysgenic crosses, always choose males when Sbq2,3 is not balanced because recombination is suppressed in males.  Also note that "yw" refers to the yellow gene and the white gene, both of which are mutant and on the X.

If the insert is on the 3rd chromosome, it is probably best to use w/w; Dr1/TMS, Sb q2,3. This cross is described last.  Note that the chromosome TMS, Sb q2,3 is a balancer so it will not recombine.

Hopping off the X Chromosome.
Cross 1. Probably best to set this up with about 40 flies total in a bottle or several vials so enough male progeny are available for cross 2.
¢¢ X[yw]/Y; Sp/SM6a; Sbq2,3/TM6Ubx  x  ¡¡ X[p(w+)]/X[p(w+)] or X[yw]; +/+; +/+ {heterozygous or homozygous insert on X chromosome}
m
Select Curly, Stubble, colored-eyed males.

Cross 2.  To insure getting several new lines, set up 50 to 100 individual crosses between individual *2,3 males and several virgin yw females.

¢¢ X[p(w+)]/Y; +/SM6a; +/ Sbq2,3  x  ¡¡ X[yw]/X[yw]; +/+; +/+ {p(w+) is somewhere in the genome of the males}
m
Select Straight-winged or curly winged, long-bristled, colored-eyed males.
{Since these have lost the X chromosome that had the original insert, p(w+) must have jumped to an autosome}

Cross 3. Mate single males to yw to establish lines with new inserts k If possible, select straight-winged, long-bristled, colored-eyed progeny and continue to propogate the line.  If the P-element hopped into Sm6a or is a 2nd chromosome homozygous lethal, eye color will segregate with curly wings.  Characterize the new lines by Southern blotting.

Hopping off 2nd Chromosome.
Cross 1.
¢¢ X[yw]/Y; Sp/SM6a; Sbq2,3/TM6Ubx  x  ¡¡ X[yw]/X[yw]; p(w+)/ p(w+) or yw; +/+ {heterozygous or homozygous insert on 2nd chromosome}
m
Select Curly, Stubble, colored-eyed males.

Cross 2.

¢¢ X[yw]/Y; p(w+)/SM6a; +/ Sbq2,3  x  ¡¡ X[yw]/X[yw]; +/+; +/+ {p(w+) is somewhere in the genome of the males}
m
Select Curly-winged, long-bristled, colored-eyed females or males

{Since these have lost the 2nd chromosome that had the original insert, p(w+) must have jumped to another chromosome.  In the case of the  males, jumps to the X will be lost.}

Cross 3.  Mate single dysgenic to yw to establish lines with new inserts k Select straight-winged, long-bristled, colored-eyed progeny and continue to propogate the line.  Characterize the new lines by Southern blotting.

Hopping off 3rd Chromosome.
Cross 1.
¢¢ X[w]/Y; +/+; Dr1/TMS,Sb q2,3  x  ¡¡ X[yw]/X[yw]; +/+; p(w+)/p(w+) or + {heterozygous or homozygous insert on 3rd chromosome}
m
Select Stubble, colored-eyed males.
Cross 2.
¢¢ X[yw]/Y; +/+; +/ TMS,Sb q2,3 x ¡¡ X[yw]/X[yw]; +/+; +/+ {p(w+) is somewhere in the genome of the male}
m
Select Stubble, colored-eye females or males.
{Since these have lost the 3rd chromosome that had the original insert, p(w+) must have jumped to another chromosome. }

Cross 3.  Mate single dysgenic to yw to establish lines with new inserts k Select long-bristled, colored-eye progeny and continue to propogate the line.  Characterize the new lines by Southern blotting.