Dataset Information

Characterization of MGC2, a Mycoplasma gallisepticum cytadhesin with homology to the Mycoplasma pneumoniae 30-kilodalton protein P30 and Mycoplasma genitalium P32.

ABSTRACT: A second cytadhesin-like protein, MGC2, was identified in the avian respiratory pathogen Mycoplasma gallisepticum. The 912-nucleotide mgc2 gene encodes a 32.6-kDa protein with 40.9 and 31.4% identity with the M. pneumoniae P30 and M. genitalium P32 cytadhesins, respectively. Functional studies with reverse transcription-PCR, immunoblotting, double-sided immunogold labeling, and attachment inhibition assays demonstrated homology to the human mycoplasmal P30 and P32 cytadhesins. These findings suggest that there is a family of cytadhesin genes conserved among pathogenic mycoplasmas infecting widely divergent hosts.

SUBMITTER: Hnatow LL

PROVIDER: S-EPMC108366 | biostudies-literature | 1998 Jul

REPOSITORIES: biostudies-literature

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Characterization of MGC2, a Mycoplasma gallisepticum cytadhesin with homology to the Mycoplasma pneumoniae 30-kilodalton protein P30 and Mycoplasma genitalium P32.

Hnatow L L LL Keeler C L CL Tessmer L L LL Czymmek K K Dohms J E JE

Infection and immunity 19980701 7

A second cytadhesin-like protein, MGC2, was identified in the avian respiratory pathogen Mycoplasma gallisepticum. The 912-nucleotide mgc2 gene encodes a 32.6-kDa protein with 40.9 and 31.4% identity with the M. pneumoniae P30 and M. genitalium P32 cytadhesins, respectively. Functional studies with reverse transcription-PCR, immunoblotting, double-sided immunogold labeling, and attachment inhibition assays demonstrated homology to the human mycoplasmal P30 and P32 cytadhesins. These findings sug ...[more]

PMID: 9632619

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Project description:The discovery of small open reading frames (smORFs) encoding for polypeptides (SEPs; <100aa) highlights that the coding capacity of the genomes has been underestimated. Most ORF-finding algorithms have historically set a minimum threshold length of 100 aa. As consequence, some transcripts encoding for SEPs, were erroneously discarded or classified as non-coding RNAs (ncRNAs). With this experiments we try to experimentally assess the existence and complexity of these small proteins. The experimental design includes: After growing each Mycoplasma for 6h at 37°C, cells were washed twice with PBS and lysed with 700 µl of Qiazol buffer. Then, samples were lysed with 700 µl of Qiazol buffer. RNA extractions were performed by using the miRNeasy mini Kit (Qiagen) following the instructions of the manufacturer. Libraries for RNA-seq were prepared following directional RNA-seq library preparation and sequencing. Briefly, 1 µg of total RNA was fragmented to ~100-150 nt using NEB Next Magnesium RNA Fragmentation Module (ref. E6150S, NEB). Treatments with Antarctic phosphatase (ref. M0289S, NEB) and PNK (ref. M0201S, NEB) were performed in order to make the 5’ and 3’ ends of the RNA available for adapter ligation. Samples were further processed using the TruSeq small RNA Sample Prep Kit (ref. RS-200-0012, Illumina) according to the manufacturer's protocol. In summary, 3’ adapters and subsequently 5’ adapters were ligated to the RNA. cDNA was synthesized using reverse transcriptase (SuperScript II, ref. 18064-014, Invitrogen) and a specific primer (RNA RT Primer) complementary to the 3’ RNA adapter. cDNA was further amplified by PCR using indexed adapters supplied in the kit. Finally, size selection of the libraries was performed using 6% Novex® TBE Gels (ref. EC6265BOX, Life Technologies). Fragments with insert sizes of 100 to 130 bp were cut from the gel, and cDNA was precipitated and eluted in 10 µl of elution buffer. Double-stranded templates were cluster amplified and sequenced on an Illumina HiSeq 2000.

Dataset Information

Characterization of MGC2, a Mycoplasma gallisepticum cytadhesin with homology to the Mycoplasma pneumoniae 30-kilodalton protein P30 and Mycoplasma genitalium P32.

Publications

Characterization of MGC2, a Mycoplasma gallisepticum cytadhesin with homology to the Mycoplasma pneumoniae 30-kilodalton protein P30 and Mycoplasma genitalium P32.

Similar Datasets

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