67
 region hybridized in Southern blots, and identifying Mspl and Ddel sites within

 each region.




 Materials and Methods

 The cloned probe pB178 was replicated in Escherichia coli strain DH5a

 (Gibco BRL) and isolated by the alkaline-lysis procedure (Sambrook, Fritsch &

 Maniatis 1989).

 Mapping restriction sites within pB178. Purified pB178 was digested

 with Pstl to release the 9.45kb honey bee DNA insert (referred to as 178),

 which was subsequently separated from the vector by electrophoresis in 0.8%

 SeaPlaque agarose (FMC) and isolated by extraction with phenol and ether.

 The relative positions of the restriction sites were determined by sequentially

 and reciprocally digesting 178 and pB178 with up to four restriction enzymes.

 pB178 was treated with one or more restriction enzymes, and one aliquot was

 set aside for analysis while another was digested with Pstl to determine the

 location of the endonuclease recognition sites relative to the Pstl ends of the

 insert. The digested DNA was electrophoresed in 1% agarose (Kodak IBI), and

 the fragments were visualized by staining with ethidium bromide. Fragment

 sizes were estimated by comparison to molecular weight standards using a HI-

 PAD digitizer. For comparison and confirmation, the fragment sizes were also

estimated using the National Center for Supercomputing Applications (NCSA)