Abstract
Abstract Conditions for efficient transformation with native and denatured T4 DNA are described. Both native and denatured DNA cotransform amH17–amN116 which spans about 7% of the T4 genome, including the complete rII cistrons, quite efficiently. Thus, r mutations induced in denatured T4 transforming DNA by heat at 100° and pH 7.1 were readily detected as r mutant transformants. The rate of heat mutagenesis from r + to r at 100° is about 0.1%/min. Native T4 DNA containing numerous single-strand nicks within the region to be donated also cotransforms amH17–amN116 . The extent of nicking in native DNA can be estimated by residual transforming activity after denaturation. Covalent closure of single-strand nicks in native DNA by DNA ligase has also been detected by denatured DNA transformation. Increases (1000-fold) in denatured DNA transforming activity for amH17–amN116 were obtained after treating highly nicked native DNA with saturating levels of T4 DNA ligase. The biological activity of small fragments of denatured T4 DNA containing no transforming activity for amH17–amN116 was restored upon renaturation. Even renatured DNA species which required two renaturation events to restore activity, and thus, contained at least three individual fragments of DNA, cotransformed amH17–amN116 and donated the complete rII cistrons efficiently.
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