Gene probes to detect cross-culture contamination in hormone producing cell lines

Summary Cross-culture contamination of cell lines propagated in continuous culture is a frequent event and particularly difficult to resolve in cells expressing similar phenotypes. We demonstrate that DNA-DNA hybridization to blotted endonuclease-digested cell DNA effectively detects cross-culture contamination to monitor inter-species as well as intra-species cross contamination. An insulin-producing cell-line, Clone-16, originally cloned from a human fetal endocrine pancreatic cell line did not produce human c-peptide as anticipated. DNA from these cells showed no hybridization to the human ALU sequence probe, BLUR, and lacked restriction fragment length polymorphism typical for the human HLA-DQ β-chain gene. Although a human insulin gene probe showed a weak, nonhuman hybridization pattern, a cDNA probe for the Syrian hamster insulin gene hybridized strongly consistent with a single copy hamster insulin gene. Karyotyping confirmed the absence of human chromosomes in the Clone-16 cells while sizes, centromere indices, and banding patterns were identical to Syrian hamster fibroblasts. We conclude that the insulin-producing Clone-16 cells are of Syrian hamster origin and demonstrate the effective use of gene probes to control the origin of cell cultures. This paper is dedicated to the late Lis Lyngsie in much appreciation of her contributions to this study. This work was supported in part by the National Institutes of Health, Bethesda, MD (grants DK 26190 and 33873). I. Matsuba and B. Michelsen were supported by research fellowships from the Juvenile Diabetes Foundation International, and J. Scholler from the Danish Medical Research Council (J.no. 12-5758).     

The role of mycorrhiza in plant populations and communities

Summary There is a large body of literature concerning the value of mycorrhiza to plant growth. Recent emphasis on the potential benefits of the fungi to natural ecosystems conservation and productive agriculture has focused attention on the roles and underlying mechanisms of the association. In parallel, recognition that isolates/ species of vesicular arbuscular mycorrhizal fungi have variable life-history traits has resulted in investigations focused on the symbiotic fungi rather than the host, a perspective which may reflect the fungal evolutionary strategies. This paper discusses progress in understanding interactions amongst hyphae, which in one form or another are a major component of mycorrhiza and each phase of the life-history.     

Formation of a minichromosome by excision of the proximal region of 17q in a patient with von Recklinghausen neurofibromatosis

An interstitial deletion, 17cen----q11.2 (or q12), and a small extra chromosome was found in a sporadic case of von Recklinghausen neurofibromatosis (NF1). In situ hybridization with a chromosome 17-specific alpha-satellite probe showed that the small chromosome was derived from the deleted region, most likely by an excision/ring formation. This chromosome rearrangement is in agreement with the localization of the von Recklinghausen neurofibromatosis (NF1) locus to the proximal region of 17q, but with a more distal breakpoint than observed in two previously described reciprocal translocations associated with NF1. If the NF1 gene has been truncated by the present rearrangement, it may suggest that the NF1 gene is a very large gene at the genomic level. Alternatively, NF1 in this patient may be caused by the gradual loss in somatic cells of the small chromosome carrying an intact NF1 gene, thereby suggesting a recessive mechanism at the gene level. Finally, an intact NF1 gene may have been placed in close proximity with alpha-satellite sequences, which might cause inactivation of the gene. The small supernumerary chromosome may not only facilitate the cloning of the NF1 gene itself, but also offers explanations of the mechanism underlying development of the disease.     

A 2.5-Mb physical map within 3p21.1 spans the breakpoint associated with Greig cephalopolysyndactyly syndrome.

Numerous investigations suggest that one or more genes residing in the p14 to p21 region of human chromosome 3 are critical to the development of neoplastic diseases such as renal cell carcinoma and small-cell lung cancer (SCLC). This region is additionally involved in several interchromosomal translocations, one of which is associated with the developmental disorder Greig cephalopolysyndactyly syndrome. A series of five loci that map in close proximity to the Greig syndrome breakpoint [t(3;7)(p21.1;p13)] at 3p21.1 have been physically linked by pulsed-field gel analysis over a 2.5-Mb region. The probes include ACY1, cA84 (D3S92), cA199 (D3S93), pHF12-32 (D3S2), and MW-Not153 (D3S332). The Greig 3;7 translocation breakpoint was discovered between clones cA199 and MW-Not153, separated by 825 kb. Further analysis revealed comigration of a rearranged fragment detected by MW-Not153 and a chromosome 7 probe previously shown to be in close proximity to the breakpoint (CRI-R944). This latter probe also detects a rearrangement in a second Greig-associated translocation, (6;7)(q27;p13). The physical map resulting from this analysis orders the markers along the chromosome and identifies several locations for CpG islands, likely associated with genes. Although probe pEFD145.1 (D3S32) has been genetically linked to D3S2 (2 cM), physical linkage to the other five loci could not be demonstrated. One of the linked loci, D3S2, has been widely utilized in the analysis of chromosome 3p loss in several malignant diseases. Since expression of ACY1, a housekeeping gene, is specifically reduced in many cases of SCLC, knowledge of its precise chromosomal position and identification of neighboring putative gene loci should facilitate investigation into the mechanism of this reduction.