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871.
Tomoko Okazaki Junko Takita Takashi Kohno Hiroshi Handa J. Yokota 《Human genetics》1996,98(3):253-258
The arbitrarily primed-PCR (AP-PCR) genomic fingerprinting method was applied to evaluate its effectiveness in detecting
and characterizing amplified DNA fragments in two small-cell lung carcinoma (SCLC) cell lines, NCI-H69 and NCI-H82. Of the
2428 DNA fragments detected by AP-PCR using 62 arbitrary primers, 2 (0.08%) DNA fragments were amplified in NCI-H69 and 6
(0.25%) DNA fragments were amplified in NCI-H82. Based on these results, we estimate the total size of the amplified genomic
regions in these cell lines to be 3000 megabase pairs (Mb) × 0.0008 = 2.4 Mb in NCI-H69 and 3000 Mb × 0.0025 = 7.5 Mb in NCI-H82.
The 2 amplified fragments in NCI-H69 were mapped to chromosome 2, and all 6 amplified fragments in NCI-H82 were mapped to
chromosome 8. This strongly suggests that restricted chromosomal regions are specifically amplified in these SCLC cell lines.
Since the N-myc gene at 2p24 is amplified in NCI-H69 and the c-myc gene at 8q24 is amplified in NCI-H82, it is possible that these DNA fragments are co-amplified with N-myc or c-myc in these cell lines. However, since the 2 amplified fragments in NCI-H69 were not amplified in 42 other human cancer cell
lines including 11 cell lines carrying amplified N-myc genes, it is also possible that there are amplified regions on chromosome 2 other than the N-myc locus at 2p24 in NCI-H69. In contrast, all 6 amplified fragments in NCI-H82 were amplified in several other human cancer
cell lines carrying amplified c-myc genes. This result further indicates that these fragments were derived from an amplification unit that includes the c-myc gene. Our results show the ability of the AP-PCR method to analyze the fraction of the genome with amplification in human
cancer cells.
Received: 10 April 1995 / Revised: 18 December 1995, 15 April 1996 相似文献
872.
Several mutant cDNAs of elongation factor 2 (EF-2) were constructed by site-directed mutagenesis and their products expressed in mouse cells were investigated. Amino acid substitution for the histidine residue of codon 715, which is modified post-translationally to diphthamide, resulted in non-functional EF-2 and this substitution did not render EF-2 resistant to Pseudomonas aeruginosa exotoxin A, which inactivates EF-2 transferring ADP-ribose to the diphthamide residue. These non-functional EF-2s with replacements of the histidine-715 residue showed various extents of inhibition of protein synthesis by competing with functional EF-2 in vivo. These results suggest that histidine-715 is essential for the translocase activity of EF-2 and that the region around diphthamide functions in recognition of, and/or binding to ribosomes. Substitution of proline for the alanine-713 residue and substitution of glutamine for the glycine-717 residue converted EF-2 to partially toxin-resistant forms. Two-dimensional gel analysis with fragment A of diphtheria toxin of these toxin-resistant EF-2s revealed that their ADP-ribosylations by toxin were much less than that of wild-type EF-2. 相似文献