Restriction digests of the clone give the following sizes (kb): BamHI--4.6; EcoRI--4.6; SalI--4.6; SalI/EcoRI--4.4, 0.3. Generic chromosome fragmentation vector containing promoters for in vitro RNA synthesis. The unique cloning sites can be used to insert targeting sequences, such as cDNAs. Any unique restriction site between the targeting sequence and the telomere-adjacent sequence can be used to linearize the plasmid before transformation. The EcoRV and ClaI sites between the SP6 promoter and the telomere sequence can be used for recovery from deletion derivatives of YAC insert sequences adjacent to the fragmentation site. A 0.24 kb EcoRI/SalI fragment containing the telomere-adjacent sequence from YTCA-1x was ligated into the MCS of pSP73. The EcoRI and BamHI sites of the MCS were destroyed and a 0.3 kb NdeI/PvuII fragment was inserted into the PvuII site of pRS303. The order of the major features in this plasmid is: bla - HIS3 - lacZ'/MCS - HindIII - SphI - PstI - AccI - SalI - TEL - ClaI - EcoRV - SP6 promoter. |
