Fig. 4.
Fig. 4. Proviral integration patterns in mice with. / BCR/ABL-induced CML-like leukemia. The gDNA was prepared from hematopoietic cells from mice with CML-like disease, digested with BglII, transferred to nylon membranes, and hybridized with a radioactive probe from the neomycin resistance gene, as described.6 Sizes (in kilobases) of DNA markers are shown at the left. (A) Balb/c mice. The gDNA from peripheral blood or spleen myeloid cells was isolated from a typical wild-type recipient ofBCR/ABL-transduced wild-type marrow (WT to WT); 2 differentGm-csf-−/− recipients ofBCR/ABL-transduced Gm-csf −/−marrow (Gm− to Gm−); 2 differentIl-3−/− recipients ofBCR/ABL-transduced Il-3−/− marrow, and 2 different double-knockout recipients of transduced double-knockout marrow (2ko to 2ko), one of which received 1 × 106 donor cells (left lane), the other of which received 0.5 × 106 donor cells (right lane). A control DNA sample containing a single proviral copy per diploid genome is included at the far right (control). The positions of bands derived from the neomycin resistance gene present in the mutated Gm-csfand Il-3 loci are indicated by the arrowheads at left. (B,C) C57Bl/6 wild-type mice. (B) The gDNA from myeloid cells from 3 representative C57Bl/6 recipients of BCR/ABL-transduced syngeneic marrow. For comparison, gDNA from leukemic cells of a Balb/c recipient of syngeneic marrow transduced with the sameBCR/ABL retroviral stock is shown at right (Balb/c), along with single copy control DNA (control). (C) The cells initiating CML-like disease in C57Bl/6 mice have multilineage repopulating ability. The gDNA from purified peritoneal macrophages (perit. Mφ), peripheral blood neutrophils (p. blood), total, B220+, and B220− spleen cells, and total, TER-119+, and TER-119− liver cells was digested with BglII and hybridized with a radioactive neo gene probe to determine the number of distinct proviral integrations in each sample (top panels). Single proviral copy control DNA from a different lane of the same blot is included at the right. The blot was stripped and reprobed with an ABL probe that detects a common 2.2-kb fragment from all proviruses as well as the endogenous murine c-abl gene (bottom panels), allowing quantitation of the proviral copy number per diploid genome.6 The same 4 proviral clones (2 major and 2 minor) are found in all hematopoietic lineages of this animal at about one copy per cell, implicating a primitive multipotential target cell, similar to that observed in Balb/c mice.6

Proviral integration patterns in mice with

BCR/ABL-induced CML-like leukemia. The gDNA was prepared from hematopoietic cells from mice with CML-like disease, digested with BglII, transferred to nylon membranes, and hybridized with a radioactive probe from the neomycin resistance gene, as described.6 Sizes (in kilobases) of DNA markers are shown at the left. (A) Balb/c mice. The gDNA from peripheral blood or spleen myeloid cells was isolated from a typical wild-type recipient ofBCR/ABL-transduced wild-type marrow (WT to WT); 2 differentGm-csf-−/− recipients ofBCR/ABL-transduced Gm-csf−/−marrow (Gm− to Gm−); 2 differentIl-3−/− recipients ofBCR/ABL-transduced Il-3−/− marrow, and 2 different double-knockout recipients of transduced double-knockout marrow (2ko to 2ko), one of which received 1 × 106 donor cells (left lane), the other of which received 0.5 × 106 donor cells (right lane). A control DNA sample containing a single proviral copy per diploid genome is included at the far right (control). The positions of bands derived from the neomycin resistance gene present in the mutated Gm-csfand Il-3 loci are indicated by the arrowheads at left. (B,C) C57Bl/6 wild-type mice. (B) The gDNA from myeloid cells from 3 representative C57Bl/6 recipients of BCR/ABL-transduced syngeneic marrow. For comparison, gDNA from leukemic cells of a Balb/c recipient of syngeneic marrow transduced with the sameBCR/ABL retroviral stock is shown at right (Balb/c), along with single copy control DNA (control). (C) The cells initiating CML-like disease in C57Bl/6 mice have multilineage repopulating ability. The gDNA from purified peritoneal macrophages (perit. Mφ), peripheral blood neutrophils (p. blood), total, B220+, and B220− spleen cells, and total, TER-119+, and TER-119− liver cells was digested with BglII and hybridized with a radioactive neo gene probe to determine the number of distinct proviral integrations in each sample (top panels). Single proviral copy control DNA from a different lane of the same blot is included at the right. The blot was stripped and reprobed with an ABL probe that detects a common 2.2-kb fragment from all proviruses as well as the endogenous murine c-abl gene (bottom panels), allowing quantitation of the proviral copy number per diploid genome.6 The same 4 proviral clones (2 major and 2 minor) are found in all hematopoietic lineages of this animal at about one copy per cell, implicating a primitive multipotential target cell, similar to that observed in Balb/c mice.6 

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