Dataset Information

Artiodactyl retroposons: association with microsatellites and use in SINEmorph detection by PCR.

ABSTRACT: During a search of polymorphic microsatellites for bovine genome mapping, we found that microsatellites often occur as tails of artiodactyl C-A retroposon elements. In this element, C (85bp) is a tRNA derivative, while A (117bp) is of unknown origin. The A element also occurs as dimer element with a connecting 27bp linker sequence comprising hexanucleotide CACTTT repeats. In 10 clones (45% of those selected deliberately for dinucleotide repeats), the microsatellite motif is associated with the C-A retroposon. In 50% of 44 database artiodactyl C-A sequences, the element also has a microsatellite tail. The microsatellite is usually a simple (CA)n repeat, but in some cases it is an apparent derivative of the linker sequence CACTTT. All but one of 33 database dimer elements have trinucleotide repeat tails (AGC)n, n = 1-9. Microsatellites, retroposons, and their truncated versions (C and/or A) often occur as clusters. We derived the consensus sequence (202bp) of the C-A element, and designed four primers for inter-SINE amplification with the aim of finding SINEmorph polymorphisms. The method is potentially powerful for rapidly producing polymorphic markers for artiodactyl genome mapping.

SUBMITTER: Kaukinen J

PROVIDER: S-EPMC312422 | biostudies-other | 1992 Jun

REPOSITORIES: biostudies-other

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Artiodactyl retroposons: association with microsatellites and use in SINEmorph detection by PCR.

Kaukinen J J Varvio S L SL

Nucleic acids research 19920601 12

During a search of polymorphic microsatellites for bovine genome mapping, we found that microsatellites often occur as tails of artiodactyl C-A retroposon elements. In this element, C (85bp) is a tRNA derivative, while A (117bp) is of unknown origin. The A element also occurs as dimer element with a connecting 27bp linker sequence comprising hexanucleotide CACTTT repeats. In 10 clones (45% of those selected deliberately for dinucleotide repeats), the microsatellite motif is associated with the C ...[more]

PMID: 1377817

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Project description:This study reports on the use of PCR to directly detect and distinguish Campylobacter species in bovine feces without enrichment. Inhibitors present in feces are a major obstacle to using PCR to detect microorganisms. The QIAamp DNA stool minikit was found to be an efficacious extraction method, as determined by the positive amplification of internal control DNA added to bovine feces before extraction. With nested or seminested multiplex PCR, Campylobacter coli, C. fetus, C. hyointestinalis, and C. jejuni were detected in all fecal samples inoculated at approximately 10(4) CFU g(-1), and 50 to 83% of the samples inoculated at approximately 10(3) CFU g(-1) were positive. At approximately 10(2) CFU g(-1), C. fetus, C. hyointestinalis, and C. jejuni (17 to 50% of the samples) but not C. coli were detected by PCR. From uninoculated bovine feces, a total of 198 arbitrarily selected isolates of Campylobacter were recovered on four commonly used isolation media incubated at three temperatures. The most frequently isolated taxa were C. jejuni (152 isolates) and C. lanienae (42 isolates), but isolates of C. fetus subsp. fetus, Arcobacter butzleri, and A. skirrowii also were recovered (</=2 isolates per taxon). Considerable variability was observed in the frequency of isolation of campylobacters among the four media and three incubation temperatures tested. With genus-specific primers, Campylobacter DNA was detected in 75% of the fecal samples, representing an 8% increase in sensitivity relative to that obtained with microbiological isolation across the four media and three incubation temperatures tested. With nested primers, C. jejuni and C. lanienae were detected in 25 and 67% of the samples, respectively. In no instance was DNA from either C. coli, C. fetus, or C. hyointestinalis detected in uninoculated bovine feces. PCR was more sensitive than isolation on microbiological media for detecting C. lanienae (17%) but not C. jejuni. Campylobacters are a diverse and fastidious group of bacteria, and the development of direct PCR not only will increase the understanding of Campylobacter species diversity and their frequency of occurrence in feces but also will enhance the knowledge of their role in the gastrointestinal tract of livestock and of the factors that influence shedding.

Project description:BACKGROUND: The popularity of microsatellites has greatly increased in the last decade on account of their many applications. However, little is currently understood about the factors that influence their genesis and distribution among and within species genomes. In this work, we analyzed carnivore microsatellite clones from GenBank to study their association with interspersed repeats and elucidate the role of the latter in microsatellite genesis and distribution. RESULTS: We constructed a comprehensive carnivore microsatellite database comprising 1236 clones from GenBank. Thirty-three species of 11 out of 12 carnivore families were represented, although two distantly related species, the domestic dog and cat, were clearly overrepresented. Of these clones, 330 contained tRNALys-derived SINEs and 357 contained other interspersed repeats. Our rough estimates of tRNA SINE copies per haploid genome were much higher than published ones. Our results also revealed a distinct juxtaposition of AG and A-rich repeats and tRNALys-derived SINEs suggesting their coevolution. Both microsatellites arose repeatedly in two regions of the interspersed repeat. Moreover, microsatellites associated with tRNALys-derived SINEs showed the highest complexity and less potential instability. CONCLUSION: Our results suggest that tRNALys-derived SINEs are a significant source for microsatellite generation in carnivores, especially for AG and A-rich repeat motifs. These observations indicate two modes of microsatellite generation: the expansion and variation of pre-existing tandem repeats and the conversion of sequences with high cryptic simplicity into a repeat array; mechanisms which are not specific to tRNALys-derived SINEs. Microsatellite and interspersed repeat coevolution could also explain different distribution of repeat types among and within species genomes.Finally, due to their higher complexity and lower potential informative content of microsatellites associated with tRNALys-derived SINEs, we recommend avoiding their use as genetic markers.

Dataset Information

Artiodactyl retroposons: association with microsatellites and use in SINEmorph detection by PCR.

Publications

Artiodactyl retroposons: association with microsatellites and use in SINEmorph detection by PCR.

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