Comparison of different protocols for DNA preparation and PCR amplification of mitochondrial genes of tardigrades

Schill, Ralph O.

doi:10.4081/jlimnol.2007.s1.164

Cited by 17 publications

(17 citation statements)

References 51 publications

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“…Currently, there are only four Milnesium ITS-2 sequences deposited in GenBank, and all were used for comparisons: JF951049 (Michalczyk et al, 2012a), GQ403681-2 (Schill et al, 2009), and HM150648 (Wełnicz et al, 2010). For the COI, seven of eight available sequences were compared with our material: FJ435810 (Guil and Giribet, 2008), JN664950 (Michalczyk et al, 2012a), EU244603-4 (Schill, 2007), and KP013598, 601 and 613 (Velasco-Castrillon et al, 2014;unpublished). To calculate molecular distances for the 28S rRNA, we used eight of the twelve deposited sequences: JX888541 and JX888585-7 (Adams et al, 2012;unpublished), FJ435779-80 (Guil and Giribet, 2008), and KC138808-9 (Zawierucha, 2012;unpublished).…”

Section: Genotypingmentioning

confidence: 99%

Experimental taxonomy exposes ontogenetic variability and elucidates the taxonomic value of claw configuration in MilnesiumDoyère, 1840 (Tardigrada: Eutardigrada: Apochela)

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Gąsiorek

Stec

et al. 2016

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Key words: barcoding, diagnostic key, development tracking, in vitro culture, M. berladnicorum, M. variefidum sp. nov. AbstractIn this paper we describe a new apochelan species, Milnesium variefidum sp. nov. from Scotland and provide novel morphological and molecular data for Milnesium berladnicorum Ciobanu et al., 2014. The new species differs from the most similar M. berladnicorum by the presence of developmental dimorphism in claw configuration, absent or weakly developed cuticular bars under claws I-III, a different arrangement of cuticular pseudoplates, and by differences in the sequences of three nuclear DNA fragments: 18S rRNA (p-distance: 0.6%), 28S rRNA (2.0%), ITS-2 (9.3%), and on mitochondrial gene COI (12.4%). Although ontogenetic claw configuration change was suspected to occur in some Milnesium species, we are the first to document it through the combined use of traditional, molecular and experimental methodologies. We discuss the implications of the observed phenomenon for the taxonomy of the genus and propose a new diagnostic key to all Milnesium species described up to the end of 2015. We also review other traits used for species differentiation in the genus and offer recommendations to improve the quality of future descriptions as well as suggest a need for integrative redescriptions of the known species. Finally, we propose to suppress M. dujiangensis and M. tardigradum trispinosum and suggest that M. alpigenum and M. quadrifidum are valid species that require thorough redescriptions.

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Section: Genotypingmentioning

confidence: 99%

Experimental taxonomy exposes ontogenetic variability and elucidates the taxonomic value of claw configuration in MilnesiumDoyère, 1840 (Tardigrada: Eutardigrada: Apochela)

Morek

Gąsiorek

Stec

et al. 2016

CTOZ

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“…The consequences of these limitations are evident in the taxon sampling in phylogenetic studies, with the first studies including three or four in‐group taxa, and the latter including 11 (Jørgensen and Kristensen, 2004) to 14 (Kiehl et al., 2007a). For molecular studies this is in part due to a requirement for extracting suitable quantities of DNA for polymerase chain reaction (PCR), achieved by pooling individuals (for exceptions see Blaxter et al., 2004; Guidetti et al., 2005; Nichols et al., 2006; Schill and Steinbrück, 2007;Schill, 2007; Sands et al., 2008). An unfortunate consequence of pooling morphotypes is that the included taxa are biased towards more commonly sampled taxa, and therefore cryptic diversity, which is often a motivation for lower level systematic studies, remains undetected.…”

Section: Introductionmentioning

confidence: 99%

Phylum Tardigrada: an “individual” approach

et al. 2008

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Phylum Tardigrada consists of 1000 tiny, hardy metazoan species distributed throughout terrestrial, limno-terrestrial and oceanic habitats. Their phylogenetic status has been debated, with current evidence placing them in the Ecdysozoa. Although there have been efforts to explore tardigrade phylogeny using both morphological and molecular data, limitations such as their few morphological characters and low genomic DNA concentrations have resulted in restricted taxonomic coverage. Using a protocol that allows us to identify and extract DNA from individuals, we have sequenced 18S rDNA from 343 tardigrades from across the globe. Using maximum parsimony and Bayesian analyses we have found support for dividing Order Parachela into three superfamilies and further evidence that indicates the traditional taxonomic perspective of families in the class Eutardigrada are nonmonophyletic and require re-working. It appears that conserved morphology within Tardigrada has resulted in conservative taxonomy as we have found cases of several discrete lineages grouped into single genera. Although this work substantially adds to the understanding of the evolution and taxonomy of the phylum, we highlight that inferences gained from this work are likely to be refined with the inclusion of further taxa-specifically representatives of the nine families yet to be sampled.

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“…Another population was recently discovered in Koror, Palau (Schill 2007: Macrobiotus 'richtersi group' 4). All specimens were cultured with bdelloid rotifers, and raised on green algae, as described by Schill (2007).…”

Section: Taxon Samplingmentioning

confidence: 98%

“…Genomic DNA was extracted from tardigrades according to Schill (2007), using the NucleoSpin Tissue method (Macherey-Nagel, Düren, Germany). The lysis was achieved by incubation of the specimens in a proteinase K/SDS solution at 56°C for 12 h, and subsequently at 70°C for 10 min.…”

Section: Its 2 Gene Amplification and Sequencingmentioning

confidence: 99%

Using compensatory base change analysis of internal transcribed spacer 2 secondary structures to identify three new species in Paramacrobiotus (Tardigrada)

et al. 2010

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Species within the tardigrade genus Paramacrobiotus could be distinguished via an analysis of internal transcribed spacer 2 (ITS2) secondary structures. Sequences of P. richtersi and four populations previously treated under provisional names (Paramacrobiotus 'richtersi group' 1 to 4) from different continents were determined and annotated, and their secondary structures were predicted. A tree based on a combined sequence-structure alignment was reconstructed by Neighbor-Joining. The topology obtained is consistent with a tree based on a distance matrix of compensatory base changes (CBCs) between all ITS2 sequence-structure pairs in the global multiple alignment. The CBC analysis, together with 18S rDNA sequences, physiological, biochemical and biophysical data identified three species new to science that are morphologically indistinguishable from P. richtersi. These are formally described under the names Paramacrobiotus fairbanksi sp. nov., P. kenianus sp. nov., and P. palaui sp. nov.

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Comparison of different protocols for DNA preparation and PCR amplification of mitochondrial genes of tardigrades

Abstract: ABSTRACT

Cited by 17 publications

References 51 publications

Experimental taxonomy exposes ontogenetic variability and elucidates the taxonomic value of claw configuration in MilnesiumDoyère, 1840 (Tardigrada: Eutardigrada: Apochela)

Experimental taxonomy exposes ontogenetic variability and elucidates the taxonomic value of claw configuration in MilnesiumDoyère, 1840 (Tardigrada: Eutardigrada: Apochela)

Phylum Tardigrada: an “individual” approach

Using compensatory base change analysis of internal transcribed spacer 2 secondary structures to identify three new species in Paramacrobiotus (Tardigrada)

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