Subject: Isolation of Df(2R)BSC3
Isolation of Df(2R)BSC3
Kevin Bogart and Kevin Cook
Bloomington Stock Center, Indiana University
To provide deletion coverage of the 49AB region, we induced a new
deficiency by P transposase-induced male recombination between
P{PZ}Cam03909 (48F1-2) and P{lacW}k15501 (49B3-6; revised cytological
localization estimated from mapping of P insertion to 24 nucleotides 5' of
Sin3A mRNA (Prokopenko et al. Genetics, in press)). We outline here the
steps used in generating the deficiency and its cytological characterization.
First, the P{lacW}k15501 insertion was marked with bw1 sp1. We
recovered a meiotic crossover event between the P{lacW}k15501 chromosome
from stock \#11142 and the cn1 bw1 sp1 chromosome from stock \#4455 to
create the stock P{lacW}k15501 unchk15501 bw1 sp1/SM6a.
To induce the deletion, we first crossed wgSp-1/CyO; ry506 Sb1
P{Delta2-3}99B/TM6 (stock \#2535) females to P{lacW}k15501 unchk15501
bw1 sp1/SM6a males. P{lacW}k15501 unchk15501 bw1 sp1/CyO; ry506
Sb1 P{Delta2-3}99B/+ males were crossed to cn1 P{PZ}Cam03909/CyO;
ry506 (stock \#11356) females to generate males of the genotype cn1
P{PZ}Cam03909/P{lacW}k15501 unchk15501 bw1 sp1; ry506 Sb1
P{Delta2-3}99B/ry506. Potential deletion-bearing (cn bw) or tandem
duplication-bearing (cn+ bw+) progeny were recovered as Sb+ males
from crosses of these males to cn1 bw1 (stock \#264) females and SM6a
balanced stocks were established from crosses to cn1 bw1 Kr1/SM6a,
bwk1 (stock \#3494) females.
In this screen of ~9,500 progeny, we recovered 19 potential tandem
duplications and 14 potential deletions in Sb+ males. The presence of a
P-to-P deletion or tandem duplication was determined cytologically in
polytene squash preparations of larvae from outcrossing males from a subset
of the balanced stocks to Canton S females. One of eight potential tandem
duplications tested was found to be duplicated cytologically. Five of ten
potential deletions tested were found to be deleted (and four of these five
deletions retained the miniwhite marker from P{lacW}). The remaining
cytological preparations showed no cytologically visible aberrations.
The single deletion line we retained has been named Df(2R)BSC3. It is
maintained in a stock of genotype Df(2R)BSC3, w+mC unchk15501 cn1
bw1 sp1/SM6a, bwk1. By cytological criteria, it has the breakpoints
48E12-F4;49A11-B6. The precise molecular breakpoints in the regions of the
two P insertions have not been determined; however, this deletion line
retains the P{lacW} miniwhite marker, which could be used as a molecular
marker for defining the sequence of the Df breakpoints.
Note that the P{lacW}k15501 insertion is separable from the unchk15501
mutation based on the noncomplementation of unch and Df(2R)CX1. We
confirmed the cytology of Df(2R)CX1 as 49C1-4;50C23-D2, which would leave
at least six bands between the P insertion site and the proximal deletion
breakpoint. Kania et al. (Genetics 139: 1663-1678, 1995) noted that the
unch phenotype could not be reverted by excision of the P{lacW} insertion.