16S rRNA gene sequence

The cultures were incubated for 48 hours. The above process was repeated to purify the colonies. Single colonies with diverse morphological characteristics were selected and inoculated into BHI liquid medium.

The cultures were stained with Gram staining, and the stained slides were observed under a BX50 optical microscope to record the cell morphology and arrangement. The isolates with uniform distribution, single morphology, and different colors were selected and stored at −80 ℃. The bacterial genomic DNA of the above bacteria was extracted using the Tiangen bacterial genomic DNA extraction kit.

The DNA of the qualified samples was amplified using polymerase chain reaction (PCR) for DNA fragments. The amplification and sequencing were performed using the universal primers 27F (5′-AGAGTTTGATCCTCGCTCAG-3′) and 1492R (5′-GGTTACCTTGTTACGACTT-3′) for amplification and sequencing. The PCR amplification system and conditions were referenced.

The PCR products were detected using 0.8% agarose gel electrophoresis technology. There was a bright and clear band at 1500 bp without a trailing phenomenon, indicating that the PCR products could meet the sequencing requirements. The PCR amplification products of the strains were sent to Shanghai Saiheng Biotechnology Co., Ltd. for sequencing under a low-temperature condition.