Direct Amplification of the Entire ITS Region from Poorly Preserved Plant Material Using Recombinant PCR

Blattner, Frank R.

doi:10.2144/99276st04

Cited by 298 publications

(166 citation statements)

References 12 publications

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“…ITS regions (ITS-1, 5.8S rDNA subunit, ITS-2) were amplified using a primer combination of ITSA and ITSB (Blattner, 1999). Each 20-µL polymerase chain reaction (PCR) contained 1.5 mM MgCl 2 , 0.2 mM of each dNTP, 0.4 µM of each primer, 1 U of Taq DNA polymerase (PanVera, Madison, WI, USA) with supplied reaction buffer at 1X concentration, and 45 ng of template DNA.…”

Section: Its Analysismentioning

confidence: 99%

Testing the utility of matK and ITS DNA regions for discrimination of Allium species

İpek¹,

İpek²,

Simon³

2014

Turk J Bot

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IntroductionAbout 750 species have been recognized within the genus Allium L. (Stearn, 1992) and at least 20 of these species are edible (Fritsch and Friesen, 2002). Economically imported and widely cultivated edible Allium species are onion (A. cepa L.), garlic (A. sativum L.), leek (A. porrum L.), chive (A. schoenoprasum L.), Chinese chive (A. tuberosum Rottl. ex Spreng), rakkyo (A. chinense G.Don) and Japanese bunching onion (A. fistulosum L.). Remaining edible Allium species are either cultivated locally or collected from wild for local consumption.The phylogeny of the genus Allium has been revised using several molecular approaches. The initial molecular approach for phylogenetic analyses of the genus Allium was based on restriction enzyme analysis of chloroplast DNA (Havey, 1991;von Berg et al., 1996). In these studies, the phylogenetic relationships among Allium species were found to be generally in agreement with their previous traditional taxonomic classification. Later nucleotide sequences of the some conserved DNA regions were utilized for the phylogenetic reconstruction of the genus Allium. The nucleotide sequence of the nuclear ribosomal internal transcribed spacer region (ITS) was one of the most commonly used DNA regions for discrimination of the

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Section: Its Analysismentioning

confidence: 99%

Testing the utility of matK and ITS DNA regions for discrimination of Allium species

İpek¹,

İpek²,

Simon³

2014

Turk J Bot

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“…The primers ITS-C and ITS-D (Blattner 1999) were modified for this study (ITS-D bryo and ITS-C bryo) and used for amplification of ITS1 and ITS2 and for sequencing reactions (Table 2). ITS 1 was amplified with primers ITS5-bryo (forward primer) and ITS-C bryo (reverse primer).…”

Section: Sampling and Outgroup Choicementioning

confidence: 99%

Genetic diversity of the relict moss Rhytidium rugosum (Hypnales) in Europe inferred from the ITS region (nrDNA)

Sabovljević

Frahm

2010

Biologia

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Post-glacial survival, potential migration routes, genetic diversity and phylogeography of the boreal moss species Rhytidium rugosum have been studied. This species is considered to be one of glacial relics of the wide but scattered Holarctic range. According to molecular data sampling from the selected European, American and Asian populations high genetic diversity of this species is present, even if this species is mostly sterile and produced sex organs extremely rarely and spread mostly asexually. Analysing the internal transcribed spacer (ITS) of the nuclear ribosomal DNA, it can be concluded that the populations of this species survived glaciations in various places in Europe and settled and re-settled present range space in various times from various refuges.

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“…To avoid false-positive scoring of individuals as females due to failures of PCR, i.e., an absence of the male-specific amplification product, we co-amplified the nuclear ribosomal DNA (nrDNA) internal transcribed spacer 2 (ITS2) together with the male-specific fragment as an internal PCR control. Therefore, we included both RnivY primers together with primers ITS-B and ITS-D (Blattner 1999) in one PCR reaction under the following cycling conditions: 4 min at 94 C followed by 35 cycles of 45 s at 92 C, 45 s at 54 C and 30 s at 72 C, and a final extension time of 10 min at 72 C. PCR products were visualised on 1.6% agarose gels stained with ethidiumbromide.…”

Section: Fragment Isolation Cloning Sequence Analysis and Scar Primmentioning

confidence: 99%

Sex-specific SCAR markers in the dioecious plant Rumex nivalis (Polygonaceae) and implications for the evolution of sex chromosomes

Stehlik

Blattner

2003

Theor Appl Genet

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We developed SCAR primers based on isolated and sequenced male-specific fragments as identified in an AFLP analysis of the dioecious plant Rumex nivalis. PCR amplification using these primers on females and males resulted in fragments exclusively present in males. Coamplification of the nuclear rDNA internal transcribed spacer 2 together with the male-specific fragment was applied as an internal control for successful PCR reactions to avoid false-negative sex scoring. With a length of about 164 bp, the AFLP fragment was of a similar size as the tandemly arranged, repetitive sequences of 180 bp located on the Y chromosomes of Rumex acetosa. The genetic distances between the Y-chromosomal sequences of R. nivalis and R. acetosa, both members of the section Acetosa, were substantial. We found intra-individual divergence among cloned sequences of the male-specific fragment in R. nivalis. The patterns of interspecific and intra-individual sequence variation found are in accordance with proposed modes of the evolution of sex chromosomes. Y chromosomes possibly arose only once in the genus Rumex and consist mainly of heterochromatic DNA. Due to the almost complete absence of selection on them, Y chromosomes are likely to accumulate large numbers of mutations.

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Direct Amplification of the Entire ITS Region from Poorly Preserved Plant Material Using Recombinant PCR

Cited by 298 publications

References 12 publications

Testing the utility of matK and ITS DNA regions for discrimination of Allium species

Testing the utility of matK and ITS DNA regions for discrimination of Allium species

Genetic diversity of the relict moss Rhytidium rugosum (Hypnales) in Europe inferred from the ITS region (nrDNA)

Sex-specific SCAR markers in the dioecious plant Rumex nivalis (Polygonaceae) and implications for the evolution of sex chromosomes

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