Additional file 1 of NADH oxidase of Mycoplasma synoviae is a potential diagnostic antigen, plasminogen/fibronectin binding protein and a putative adhesin
Creators: name: Hu, Zengjin
name: Li, Haoran
name: Zhao, Yuxin
name: Wang, Guijun
name: Shang, Yuanbing
name: Chen, Yuetong
name: Wang, Shaohui
name: Tian, Mingxing
name: Qi, Jingjing
name: Yu, Shengqing
Publisher: name: figshare
Year: 2023
Subjects: subject: Biochemistry
subject: Microbiology
subject: FOS: Biological sciences
subject: Cell Biology
subject: Molecular Biology
subject: Environmental Sciences not elsewhere classified
subject: Chemical Sciences not elsewhere classified
subject: Immunology
subject: FOS: Clinical medicine
subject: Cancer
subject: Infectious Diseases
subject: FOS: Health sciences
subject: Virology
subject: Cardiology

description: Additional file 1: Table S1. Primers used for overlap PCR amplification of MSnox [26] and MSfba  [56] genes. Fig. S1. Detection of antibody titers of different rabbit antisera by ELISA. The ELISA plates were coated with 0.5 μg of rMSNOX , rMSFBA, MS whole cell proteins or MG whole cell proteins respectively. Then the plates were reacted with serially diluted rabbit anti-rMSNOX, anti-rMSFBA, anti-MS or anti-MG serum, respectively. When the ratios of the OD450nm value of the antiserum and the pre-immune serum (marked above the black column graph) was greater than 2.1, the maximum dilution was determined as the antibody titer of the antiserum. Fig. S2. Full-length figure for expression and purification of rMSNOX and rMSFBA protein. Lane M: protein marker; lane 1 and 5: cell lysates of E. coli BL21 containing empty vector; lane 2 and 6: total cell lysates of recombinant strain E. coli BL21 (pET28a-MSnox) and E. coli BL21 (pET28a-MSnox); lane 3 and 7: supernatant of total cell lysates of recombinant bacteria E. coli BL21 (pET28a-MSnox) and E. coli BL21 (pET28a-MSnox); lane 4 and 8: purified His-tagged MSNOX protein and His-tagged MSFBA protein. Fig. S3. Original figure for immunogenicity analysis of rMSNOX protein. Lane M: protein marker; lane 1 and 3: purified His-tagged rMSFBA protein (His-tag control) reacted with rabbit anti-rMSNOX serum and pre-immune rabbit serum (1:1000), respectively; lane 2 and 4: purified His-tagged MSNOX rabbit reacted with rabbit anti-rMSNOX serum and pre-immune rabbit serum (1:1000), respectively. Fig. S4. Full-length blots for reactivity analysis of rMSNOX with different chicken sera. Purified rMSNOX protein (0.5 μg/well) was subjected to 12.5% SDS-PAGE followed by transferring to NC membrane. The NC membranes were cut to react with different chicken sera, including positive chicken serum against different MS isolates (MS WVU1853, MS JS1, MS HB1, MS SD1 and MS SH1), positive chicken sera of different MG isolates (MG Rlow, MG 08, MG 013, MG FBH, MG SGN and MG SS), positive sera against other avian pathogens (MI, E. coli O1/O2/O78, SPG, PM, STA, NDV, IBDV and IBV), three field MS-negative sera (FN-1, FN-2 and FN-3), and negative serum from SPF chicken. All of the chicken sera were diluted at 1:500. For Western blots using positive chicken serum against PM and STA, whole cell proteins of PM or STA were in lane 1 respectively, and 0.5 μg of rMSNOX protein were in lane 2. Fig. S5. Full-length figures for cPlg and hFn binding assays using Western blots. Serially diluted rMSNOX protein (lanes 1-3: 2, 1, and 0.5 μg) and 2 μg BSA (lane 4) were incubated with 10 μg/mL of cPlg (A) or hFn (B), and then recognized by rabbit anti-cPlg or anti-hFn polyclonal antibody (1:1000).