The nuclear DNA content in 10 species of chondrostean fishes was measured by flow cytometry. The sterlet Acipenser ruthenus blood cells were used as an internal standard. The sterlet DNA content was calculated on the basis of comparison with the Xenopus laeuis blood cells, 2C = 6.30 pg. In the tetraploid A. ruthenus and A. stellatus the DNA content comprises 3.74 pghucleus and is practically invariant; in Huso dauricus it is almost the same, 3.74-3.81 pg; and in A. nudiuentris it is a little higher, 3.88-4.04 pg.In the oldest chondrostean, Pseudoscaphirhynchus kaufmanni, the nuclear DNA content is slightly lower, 2C = 3.46-3.48 pg, and in the American paddlefish Polyodon spathula it is lower still, 3.17 pg. In two octoploid sturgeons, A. baeri and A. gueldenstaedti, the DNA content is twice as high as that of the sterlet, 8.29-8.31 and 7.86-7.88 pg, respectively; a very similar amount, 8.24-8.42 pg, was determined in the hybrid Huso huso x A. ruthenus. In the Sakhalin sturgeon, A. medirostris ( = A . mikadoi), the DNA content is two times higher than in the octoploids, 13.93-14.73 pg; therefore its ploidy may be 16n and the number of chromosomes could be 500. o 1993 Wiley-Liss, Inc.

A technique for DNA amount determination by flow cytometry based on the use of 7-amino-actinomycin D (7-amino-AMD), a fluorescent analogue of antibiotic actinomycin has been investigated, and a particular staining procedure has been developed. The procedure includes short fixation in 70% ethanol and staining for 20 min in 10W5M solution of 7-amino-AMD at pH7. The results of DNA content measurements are very reproducible. The histograms obtained have a coefficient of variation less than 3%.The absorption maximum of the complex of 7-amino-AMD with DNA is situated in the green spectrum region, making this stain particularly suitable for argon laser flow cytornetry.

The DNA targets may be labeled and simultaneously amplified in the polymerase chain reaction (PCR) using a pair of respective primers after elongation with nucleoside-5'-triphosphates carrying photoreactive groups. The amplified DNA may be subsequently photoactivated by irradiation above 300 nm, resulting in photo-cross-linking of the strands. For this goal 5-[3-(E)-(4-azido-2,3,5,6-tetrafluorobenzamido)propenyl-1]-, 5-{N-[N'-(4-azido-2,3,5, 6-tetrafluorobenzoyl)-3-aminopropionyl]aminomethyl}-, and 5-{N-[N'-(2-nitro-5-azidobenzoyl)-3-aminopropionyl]aminomethyl}-2'-de oxyuridine-5'-triphosphate (VII, VIa, and VIb) derivatives have been synthesized. It was found that VII is capable of efficiently elongating DNA primers with both Klenow fragment DNA polymerase I and Thermus aquaticus DNA polymerase. Thereto, it turned out to provide quantitative incorporation in DNA as revealed by the formation of the full-length amplificate by PCR in the presence of this photoreactive analogue without any dilution with natural dTTP. On the contrary, it was found, that incorporation of VIa and VIb do not permit further DNA replication.

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