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We postulated that plasmid DNA was probably supplied naturally from dead cells through cell disruption because this is the most natural manner of DNA supply in culture.
It has been suggested that these noncoding regions, previously termed "junk DNA," supply a vast array of ncRNAs which control the different epigenomes supporting development and generated throughout life in response to diet and environment.
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Hybridization was carried out in 440 μl of mixture containing 750 ng of Cy3- and Cy5-labeled cDNA probes, control targets, and the fragmented DNA supplied by the manufacturer at 60°C for 17 h.
For Asterias rubens (DNA supplied by A. van Wormhoudt, Station Marine de Concarneau), a partially amplified Amy gene was used as a probe for screening a mini-library The same method was applied to Ceratitis capitata and Apis mellifera (for the latter, a genomic library was kindly screened by M. Solignac in our laboratory).
Lambda DNA supplied by the manufacturer served as standard sample at a stock concentration of 100 ng/μl.
For each batch, the RLB detected the positive control CMhm and Mhf DNA supplied by S. Tasker, and the negative control was consistently negative.
Signals were read using an Applied Biosystems AB7000 sequence-detection system and were calculated against a dilution series of synthetic amplicon DNA supplied by Applied Biosystems and standards of B. divergens DNA (30,000, 3,000, 300 and 30 DNA copies).
A total of 4 μg of plasmid DNA was used for transfection in each well, and the total DNA supplied was kept constant in all cases by adjusting with empty vector.
This may be another possible DNA-supply mechanism in cell-to-cell transformation.
We used methylated DNA standards supplied with the kit, while for blanks we simply omitted DNA from the assay.
DNA was supplied from the Vermont cohort.
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