Methanocatella millerae SM9
Introduction
Methanocatella millerae SM9 (formerly Methanobrevibacter millerae) was isolated from the rumen of a sheep fed a fresh forage diet[1]. Although initially identified as Methanobrevibacter sp. or Methanobrevibacter smithii, SM9 shares 99% 16S rRNA gene sequence identity with the Methanocatella (Methanobrevibacter) millerae type strain ZA-10ᵀ (DSM 16643), supporting its assignment to this species[2]. Its robust growth and high plating efficiency make SM9 a strain of particular interest to the Global Methane Hub.
Global Methane Hub Metadata
| Origin: | Lucidome Bio, NZ |
| Sequenced by: | Plasmidsaurus, ONT + Illumina |
| Assembled by: | Nana Shao |
| Methylome called by: | William G. Alexander |
| Methyltransferases cloned by: | Roy Rodriguez Carrero |
| MIC and Culturing conditions by: | Nana Shao |
| Transformation by: |
Genome
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DefenseFinder Results
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Methylome
Pending Roy's input.
- No 6mA motifs present in output file, skipping...
- Searching dataset for following motifs: CCTG, CAGG, GGNCC
- Motifs explain the following about the (5mC) modified dataset:
- Low-quality methylated sites: 3603/4435 explained (81.2401352874859%)
- Mid-quality methylated sites: 12352/13030 explained (94.7966231772832%)
- High-quality methylated sites: 8337/8480 explained (98.31367924528301%)
- No 4mC motifs present in output file, skipping...
- Only one replicon present, skipping differential genomic coverage
Methylating E. coli
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Culturing Conditions
M. millerae SM9 is cultivated in Modified Methanobrevibacter smithii and Methanosphaera stadtmanae medium (MMSM) at 37°C. To ensure good gas transfer to the media, tubes are incubated on their sides (or at a shallow angle so that the media does not contact the stoppers) with gentle shaking at 50 rpm and 207 kPa of H2/CO2 (80:20, v/v) as an energy substrate for 2-4 days. This methanogen can also be cultivated on MMSM plates using 1.5% agar as a gelling agent with the same pressure of H2/CO2 as an energy substrate for 3-7 days.
Antimicrobial MIC Tests
Puromycin MIC Determination on Solid Media
Puromycin susceptibility of M. millerae SM9 was assessed on solid medium under anaerobic conditions, using the absence of visible colonies as the endpoint. The apparent MIC was 10 µg/mL at an inoculum of 10⁵ cells/plate and 20 µg/mL at 10⁷ cells/plate, demonstrating an inoculum-dependent difference in puromycin susceptibility under the tested conditions.
Puromycin MIC Determination on Liquid Media
Puromycin susceptibility of M. millerae SM9 was assessed on solid medium under anaero
| Antibiotics | Concentration | |
|---|---|---|
| Liquid | Solid | |
| Puromycin | ||
| nourseothricin | μM | |
Protocols
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Transformation Results
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External Links
- ↑ Joblin KN. (1999) Ruminal acetogens and their potential to lower ruminant methane emissions. Australian Journal of Agricultural Research 50, 1307–1314. https://doi.org/10.1071/AR99004
- ↑ Kelly WJ, Pacheco DM, Li D, Attwood GT, Altermann E, Leahy SC. The complete genome sequence of the rumen methanogen Methanobrevibacter millerae SM9. Stand Genomic Sci. 2016 Aug 17;11:49. doi: 10.1186/s40793-016-0171-9.
