TABLE 1.

Frequently used methods in feline microbiota studies.

Techniques	Purpose	Summary
FISH	Detection and quantification of bacterial cells	Fluorescent dye-labeled oligonucleotide probe hybridizes to ribosomal RNA sequence in cells fixed on slides with wells. Enumeration by epifluorescence microscopy
PCR/DGGE	Profiling the composition of bacterial communities for comparative analysis	Separation of 16S rDNA fragments from different bacterial types is based on differences in chemical stability, through a linearly increasing gradient of chemical denaturants. The profile of DNA fragments represents the genetic fingerprint of the community.
Qpcr	Quantification of bacteria	PCR primers and a labeled probe (often incorporating a reporter dye and a quencher molecule) are used to measure the real-time accumulation of a specific target sequence
454-Pyrosequencing	Detecting the nucleotide incorporated	A single strand of DNA is used as a template to synthesize the sequence of its complementary strand, which is determined by a chain of reactions resulting in light being emitted when a specific nucleotide or length of nucleotides are added to the complementary sequence
Shotgun sequencing	Determining the sequence of entire chromosomes and genomes	Based on producing random fragments of DNA that are then assembled by computers that order fragments by finding overlapping ends

FISH, fluorescence in situ hybridization; PCR/DGGE, polymerase chain reaction combined with denaturing gradient gel electrophoresis; qPCR, real-time quantitative polymerase chain reaction. Summarized from Tannock (2005), Suchodolski et al. (2009, 2015), Suchodolski (2011b), and Swanson et al. (2011).