Cloning of citrate permease gene of Lactococcus lactis subsp. lactis biovar diacetylactis NIAI N-7 and expression in citrate-negative lactococci.

摘要

The putative citrate permease (Cit-P) plasmid DNA (pN7CP) from L. lactis subsp. lactis biovar diacetylactis (L. diacetylactis) NIAI N-7 was cloned. Cit-P+ transformants possessed pIL253 with a 3.8-kb (pKAK1) or a 2.0-kb fragment (pKAK7). No Xba I site was present in the 3.8-kb of the Xba I-Xba I fragment from pN7CP. It may thus be possible to obtain the 2.0-kb fragment by deleting the 3.8-kb fragment. Assessment was made of citrate-transporting activity in transformants KAK1 and KAK7. Several Cit-lactococci were transformed by pKAK7. All their transformants except KK-7LM and KK-7SL of L. lactis subsp. lactis LM0230 and NIAI SLN were incapable of transporting citrate. KK-7LM and KK-7SL showed the Cit+ phenotype and citrate-transporting activity. L. diacetylactis and L. lactis subsp. lactis were classified under citrate metabolism. However, the present findings strongly indicate some strains of L. lactis subsp. lactis to be the original citrate utilizers but to have lost the Cit-P gene.