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Porcine dorfin: molecular cloning of the RNF19 gene, sequence comparison, mapping and expression analysis

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Abstract

Dorfin, encoded by the RNF19 gene, is a protein containing two RING finger motifs. Dorfin functions as an E3 ubiquitin ligase that interacts with UBE2L3/UBCH7 and UBE2E2/UBCH8, but not other ubiquitin-conjugating enzymes. Dorfin is found expressed in Lewy bodies, neuronal protein inclusions occurring in Parkinson’s disease brains. This work reports the cloning and analysis of the porcine (Sus scrofa) homologue of dorfin. The RNF19 cDNA encoding dorfin was amplified by reverse transcriptase polymerase chain reaction (RT-PCR) using oligonucleotide primers derived from in silico sequences. The porcine RNF19 cDNA codes for a protein of 838 amino acids which shows a very high similarity to human (97 %) and mouse (93 %) dorfin. The genomic organization of the porcine RNF19 gene is very similar to its human counterpart. Expression analysis by RT-PCR demonstrated that the porcine RNF19 transcript was observed in all organs and tissues examined, although differentially expressed. The highest expression of RNF19 mRNA was observed in cerebellum, heart, frontal cortex and muscle. RNF19 transcript was detected as early as 60 days of gestation in many different brain areas. Radiation hybrid mapping data indicate that the porcine RNF19 gene maps to chromosome 4 (4p11–p12). This particular map location is fully consistent with the currently known conservation of genome organization between human and pig and provides further confirmation that we have characterized the porcine homologue of the human RNF19.

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Abbreviations

bp:

Base pair

EST:

Expressed sequence tag

GAPDH:

Glyceraldehyde 3-phosphate dehydrogenase

kb:

Kilobases

ORF:

Open reading frame

PCR:

Polymerase chain reaction

PD:

Parkinson’s disease

pI:

Isoelectric point

RT-PCR:

Reverse transcription PCR

SNP:

Single nucleotide polymorphism

UTR:

Untranslated region

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Acknowledgments

We gratefully acknowledge Dr. Martine Yerle of INRA Toulouse, France for providing the pig-rodent hybrid panel. The authors wish to thank Connie Jakobsen Juhl for excellent technical assistance, and Ms. Janne Hansen for critical reading of the manuscript.

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Authors and Affiliations

  1. Department of Molecular Biology and Genetics, Science and Technology, Aarhus University, P.O. Box 50, 8830, Tjele, Denmark

    Knud Larsen, Lone Bruhn Madsen & Christian Bendixen

Authors
  1. Knud Larsen
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  2. Lone Bruhn Madsen
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  3. Christian Bendixen
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Corresponding author

Correspondence to Knud Larsen.

Additional information

The sequence of the porcine RNF19 cDNA, encoding the dorfin protein, and the genomic sequences of RNF19 have been submitted to DDBJ/EMBL/GenBank under the accessions numbers DQ090004, DQ091175, DQ091176, JQ031558 and JQ063125, respectively.

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Larsen, K., Madsen, L.B. & Bendixen, C. Porcine dorfin: molecular cloning of the RNF19 gene, sequence comparison, mapping and expression analysis. Mol Biol Rep 39, 10053–10062 (2012). https://doi.org/10.1007/s11033-012-1874-7

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  • DOI: https://doi.org/10.1007/s11033-012-1874-7

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