G0885

Unlabeled standards were prepared for all relevant analytes, and were added directly to the biomass pellet prior to the execution of the protocols described below. For amino acids, 40 µL of a 2.5 mM per amino acid solution (Pierce 20088) was used. For RNA, a 1 mg/mL solution (ribonucleic acid from torula yeast, Sigma R6625) in water was prepared, of which 80 µL was added to the sample. For glycogen, a 0.1 mg/mL solution (glycogen from bovine liver, Sigma G0885) in water was prepared, of which 100 µL was added to the sample. For fatty acids, a solution of 0.3 mg/mL of myristic acid (C14:0), palmitic acid (C16:0), palmitoleic acid (C16:1), stearic acid (C18:0), and elaidic acid (C18:1) in hexane was prepared, of which 20 µL was added.

Lactobacillus casei var rhamnosus GR-1 (L. Rhamn. ATCC 55826) was used in this study. The nucleotide sequence of HIV gp41-64 and anti-erythromycin were cloned into the L. rhamn genome. L. rhamn was cultured using Man Rigosa Sharpe broth (Sigma Aldrich, Cat# 69966) containing 32μg/ml erythromycin. After seeding, the culture tubes were sealed and cultured without shaking at 30°C for 48 hrs. At the end of culture, the L. rhamn were washed three times with DPBS before being resuspended in 20mg/ml glycogen (Sigma Aldrich G0885) solution, and stored at 4°C overnight prior to inoculation. The colony forming units were determined on Bromo Cresol Purple agar (Himedia labs M1351) plated containing 30μg/ml erythromycin, and titrated generally around 1–2×10¹⁰ per ml.