Selenoprotein A of the glycine reductase complex from Clostridium sticklandii has been purified and partially sequenced around the selenocysteine residue. An oligonucleotide probe to this region has been synthesized and shown by Southern blot analysis to hybridize to a single 2300 base pair (bp) fragment of clostridial DNA digested to completion with Hind III. A clostridial gene library of 1500 to 3000 bp DNA fragments from a complete digestion with Hind III was constructed in pUC-13 and cloned into Escherichia coli strain DH5 . Fifteen clones were selected by colony hybridization to the probe. Restriction enzyme digestion analysis of the selected clones revealed that each clone contains the same 2300 bp clostridial DNA fragment. The fragment was digested with restriction enzyme TaqI to yield four smaller fragments. A 400 bp fragment was found to hybridize to the probe. It has been subcloned into the restriction enzyme AccI site of bacteriophage M13 for the production of a single-stranded template for DNA sequencing.
