HIGHLY UNSTABLE (GATA)N-CONTAINING SEQUENCES OF THE MOUSE DURING THE CLONING PROCESS

Mouse DNA fingerprints were obtained by HaeIII digestion of genomic DNA and in-gel hybridization with the (GATA)4 oligonucleotide probe. In order to obtain locus-specific probes that hybridize with only one fragment of the (GATA)4 DNA fingerprint, a genomic library of size-selected inserts was constructed using a system of direct subcloning from the phage clones. During the cloning procedure, the phage as well as the plasmid insert DNAs changed primarily within their repetitive DNA but also within adjacent nonrepetitive sequences, as was demonstrated for several clones by in-gel hybridization with the (GATA)4 probe as well as by sequence analysis. Isolated subclones varied within their (GATA)n repeats, resulting in different insert lengths. Several "metastable" as well as stable (GATA)4-positive subclones could be isolated. Also, vector sequences were affected by alterations during the cloning process. These phenomena are discussed within the context of possible mechanisms for cloning artifacts.