Thesis (Ph. D.)--University of Wisconsin--Madison, 1949. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 136-145).

Using proteinase negative variants of Streptococcus cremoris UC310 or UC320 to manufacture cottage cheese, theoretical yields were increased 1 .97% and 1 .56% respectively when compared to the theoretical yields of the proteinase positive parents. Yield differences were strain dependant and differences between positive and negative variants were not manifest with strains of UC73 and UC97. It was necessary to produce bulk culture using pH control and to add sufficient nitrogenous stimulant to provide carry-over stimulant into the cheese milk. All cultures examined developed normally even when the bulk medium contained a blend of 5% yeast extract and casein hydrolysate. It was possible to use the culture to complete the required acidification after direct acidification to pH 5.2 with phosphoric acid. Careful selection of lactic

The effect of proteolytic enzymes on the transformation and transfection of Streptococcus lactis LM2301 protoplasts was examined in an attempt to eliminate the variability observed. By using both chymotrypsin and mutanolysin to form protoplasts followed by spread plating, consistent frequencies of 104 to 105 transformants per μg of pGB301 DNA, and 105 transfectants per μg of c2 bacteriophage DNA where achieved. Optimum transformation and transfection frequencies were obtained when 16 h cultures were treated simultaneously with 25 U/ml mutanolysin and 1.25 U/ml chymotrypsin for 15 min. Trypsin and pronase in conjunction with mutanolysin also increased transformation frequencies higher than when mutanolysin was used alone, but pronase was not as effective as chymotrypsin or trypsin. These results may explain the variability in transformation of mutanolysin-treated cells of S. lactis since commercial sources of mutanolysin contain varying amounts of proteolytic enzyme activity. Transformation of Streptococcus cremoris CS224 at low frequency (5 transformants per μg of pGB301 DNA) was achieved. Plasmid pGB301 was able to replicate and express antibiotic resistance in the resultant transformant (designated S. cremoris SW301). The presence of pGB301 in S. cremoris SW301 was confirmed by agarose gel electrophoresis.

Whole milks were adjusted to pH 5.8, 6.2, or 6. 7 with HCl and batch pasteurized at 63°C for 30 min. Each was concentrated 5:1 (40% total solids) through a single tube polysulfone membrane Abcor ultrafiltration unit. Lactose (L), casein hydrolysate (CH), and one of two brands of yeast extract (YE1, YE2) were added into cooled retentates at 0.1, 0.3, 0.5, 0. 7 or 0.9% and equilibrated overnight at 4°C. Five percent proteinase positive (Prt+) Streptococcus cremoris UC 310+ (v/w) milk based culture was added. Unfortified retentate was also inoculated with 0.1, 0.3, 0.5, 0. 7 or 0.9% starter and pH readings were taken on all samples for 24 h during incubation at 23°C. Similar substrates were inoculated with proteinase negative (Prt-) S. cremoris UC 310-. Lactose had no significant effect on acid production. Casein hydrolysate had a slight positive effect. Yeast extract had a significant effect at all preacidification levels and a significant difference was also noticed between the brands. Mean times required for the proteinase positive culture to reach pH 5.1 in 5x retentate from milk acidified to pH 5.8 were 24, 12, 10, 10, and 24 h for L, CH, YE1, YE2, and the control respectively. Proteinase negative variants of this strain had mean times of >24 h, 14 h, 11 h, 11 h, and >24 h respectively. These time differences were significantly different between Prt+ and Prt- variants. A minimum concentration of 0.2% yeast extract produced the most stimulation while greater quantities provided no additional benefit. Taste panelists were unable to detect yeast extract in retentates fermented by either culture variant.