Additional file 1 of AdpA, a developmental regulator, promotes ε-poly-l-lysine biosynthesis in Streptomyces albulus
Creators: name: Huang, Rui
name: Liu, Honglu
name: Zhao, Wanwan
name: Wang, Siqi
name: Wang, Shufang
name: Cai, Jun
name: Yang, Chao
Publisher: name: figshare
Year: 2022
Subjects: subject: Genetics
subject: FOS: Biological sciences

description: Additional file 1: Figure S1. The upstream and downstream genes of different adpA genes and phylogenetic analysis of AdpA homologs. (A) Different adpA genes and their upstream and downstream genes in three Streptomyces species. The sequence identities between the encoded proteins of homologous genes with same color are indicated. (B) Phylogenetic analysis of the novel AdpASa, four heterologous AdpA homologs AdpASd, AdpA-SH, AdpASn, AdpA-C with NCBI BLASTP hits and well-studied ones. AdpASa is marked with the red hollow circle, while the four heterologous AdpA homologs AdpASd, AdpA-SH, AdpASn, AdpA-C are all marked with the blue hollow diamond. Different clades or subclades are indicated with different colors, and some bootstrap values of the phylogenetic tree are also displayed. Figure S2. Construction of S. albulus NKA and effect of AdpASa on specific ɛ-PL formation rate. (A) Schematic method for overexpressing adpASa in S. albulus. (B) Confirmation of the integration of adpASa gene into the genome of S. albulus by PCR. Lane M, DNA marker III; Lanes1-6, amplification products using primers SET-F/SET-R with gDNA from S. albulus SET, plasmid pSET152 DNA, gDNA from S. albulus NKA, plasmid pSET152-adpASa DNA, gDNA from S. albulus NK660 and ddH2O as templates, respectively. (C) Transcription levels of adpASa gene in S. albulus NK660 and S. albulus NKA by RT-qPCR analysis. ***P &lt; 0.001 (Student's t-test). Error bars stand for the SD for three biological replicates. (D) Specific formation rates of ɛ-PL in S. albulus NKA and control strains (S. albulus SET and S. albulus NK660) cultured in fermentation medium for 100 h. Error bars stand for the SD for three biological replicates. (E) Phenotypes of S. albulus NK660 and S. albulus SET grown on MSF agar plates for 4 days. Figure S3. Construction of S. albulus SDA, S. albulus SHA, S. albulus SNA and S. albulus SCA. (A) Schematic method for expressing heterologous adpA genes in S. albulus. The blue arrow stand for heterologous adpA genes (adpASd, adpA-SH, adpASn, adpA-C). (B) Confirmation of the construction of mutants expressing heterologous adpA genes by PCR. Lane M, DNA marker III; Lanes1, 3, 5, 7, 9, 11, amplification products using gDNA from S. albulus SET, S. albulus SDA, S. albulus SHA, S. albulus SNA, S. albulus SCA and S. albulus NK660 as templates, respectively; Lanes 2, 4, 6, 8, 10, 12, amplification products using plasmid pSET152, pSET152-adpASd, pSET152-adpA-SH, pSET152-adpASn, pSET152-adpA-C and ddH2O as templates, respectively. And all the amplification products use the primers SET-F/SET-R. (C) RT-PCR results among S. albulus NK660, S. albulus SET and the four heterologous adpA genes expression mutants. Lane M, DNA marker III; Lanes D1-D10, amplification products using primers DadpA-F/DadpA-R with RNA from S. albulus NK660, cDNA from S. albulus NK660, RNA from S. albulus SET, cDNA from S. albulus SET, RNA from S. albulus SDA, cDNA from S. albulus SDA, gDNA from S. albulus SDA and ddH2O as templates, respectively. Lanes H1-H10, Lanes N1-N10 and Lanes C1-C10 are the amplification products respectively using primers HadpA-F2/HadpA-R, NadpA-BF/NadpA-BCR and CadpA-F/CadpA-R with the same templates and template orders as Lanes D1-D10. Table S1. Primers used in this study. Table S2. Accession number and length of the whole candidates used for phylogenetic analysis.