Saline One Month Mouse 602
Three-month-old female C57BL6/J mice (Jackson Laboratory) were group housed 2-3 per cage under a constant 12-hour light:12-hour dark cycle in a climate-controlled environment. Food and water were provided ad libitum. Mice in the same treatment groups were housed together. All procedures were approved by the Institutional Animal Care and Use Committee at the University of Arkansas for Medical Sciences (UAMS; Little Rock, Arkansas). Cyclophosphamide, methotrexate, and fluorouracil were purchased from the UAMS Inpatient Pharmacy. For 4 weeks, mice received weekly intraperitoneal (IP) injections on days 1, 8, 15, and 22. The control group received saline (0.9% sodium chloride) and the CMF group received 60 mg/kg cyclophosphamide, 4 mg/kg methotrexate, and 60 mg/kg fluorouracil (Anderson et al.). Drugs were diluted with sterile saline and stored per the manufacturer instructions. Drugs were mixed immediately prior to injections. Mouse fecal pellets were sent to RTL Genomics (Lubbock, Texas) for DNA extraction, amplification, and sequencing of the V3-V4 small ribosomal subunit (16S rRNA) hypervariable region about a month after treatment.