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      Isolation and characterization of microsatellite markers for Hypochaeris incana (Asteraceae) and close relatives 1

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          Abstract

          Premise of the study:

          We developed microsatellite markers to study clonal growth and interspecific hybridization in the Patagonian and subantarctic plant Hypochaeris incana (Asteraceae) and its closest relatives.

          Methods and Results:

          We developed primers for microsatellite loci from 454 sequence reads of genomic DNA of H. incana. We tested them on individuals of H. acaulis, H. hookeri, H. incana, H. palustris, and H. tenuifolia. We selected 15 polymorphic microsatellite loci, which delivered clearly scorable fragments in most or all species. With mean values between 0.7 and 0.8, the expected heterozygosity in populations of H. incana is high.

          Conclusions:

          Due to high levels of polymorphism, the developed markers make it possible to distinguish between genets and ramets in H. incana. In some markers, null alleles complicate the scoring of genotypes in tetraploids. All of the developed markers are suitable to study interspecific hybridization among this group of closely related species.

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          Most cited references8

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          msatcommander: detection of microsatellite repeat arrays and automated, locus-specific primer design.

          msatcommander is a platform-independent program designed to search for microsatellite arrays, design primers, and tag primers using an automated routine. msatcommander accepts as input DNA sequence data in single-sequence or concatenated, fasta-formatted files. Search data and locus-specific primers are written to comma-separated value files for subsequent use in spreadsheet or database programs. Binary versions of the graphical interface for msatcommander are available for Apple OS X and Windows XP. Users of other operating systems may run the graphical interface version using the available source code, provided their environment supports at least Python 2.4, Biopython 1.43, and wxPython 2.8. msatcommander is available from http://code.google.com/p/msatcommander/. © 2007 The Author.
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            Modulation of non-templated nucleotide addition by Taq DNA polymerase: primer modifications that facilitate genotyping.

            Taq DNA polymerase can catalyze non-templated addition of a nucleotide (principally adenosine) to the 3' end of PCR-amplified products. Recently, we showed that this activity, which is primer-specific, presents a potential source of error in genotyping studies based on the use of short tandem repeat (STR) markers. Furthermore, in reviewing our data, we found that non-templated nucleotide addition adjacent to a 3' terminal C is favored and that addition adjacent to a 3' terminal A is not. It was clear, however, that features of the template in addition to the 3' terminal base also affect the fraction of product adenylated. To define consensus sequences that promote or inhibit product adenylation, we transplanted sequences between the 5' ends of the reverse primers of markers that are adenylated and those of markers that are not adenylated. It proved difficult to identify a single sequence capable of protecting the products of all markers from non-templated addition of nucleotide. On the other hand, placing the sequence GTTTCTT on the 5' end of reverse primers resulted in nearly 100% adenylation of the 3' end of the forward strand. This modification or related ones (called "PIG-tailing") should facilitate accurate genotyping and efficient T/A cloning.
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              Genetic differentiation between individuals

              Rousset (2000)
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                Author and article information

                Journal
                Appl Plant Sci
                Appl Plant Sci
                apps
                Applications in Plant Sciences
                Botanical Society of America
                2168-0450
                October 2017
                19 October 2017
                : 5
                : 10
                : apps.1700081
                Affiliations
                [2 ]Institute of Botany, Department of Integrative Biology and Biodiversity Research, University of Natural Resources and Life Sciences, Vienna, Gregor-Mendel-Straße 33, 1180 Vienna, Austria
                [3 ]Instituto de Botánica Darwinion, Labardén 200, Casilla de Correo 22, B1642HYD San Isidro, Buenos Aires, Argentina
                Author notes
                [1]

                The authors thank all collectors of plant material; A. Calvo (Bariloche) for permission to collect on his property; and J. Böckelmann, C. König (both Vienna), and A. López (San Isidro) for help with flow cytometric measurements and marker development. Financial support was provided by the Ministerio de Ciencia, Tecnología e Innovación Productiva (MINCyT, Argentina; project AU/10/16 to E.U.), the Österreichische Austauschdienst (OeAD, Austria; project AR 27/2011 to K.T.), a Eurasia-Pacific Uninet scholarship to P.W., and the University of Natural Resources and Life Sciences, Vienna.

                [4]

                Present address: College of Forestry, Northwest A&F University, Taicheng 3, Yangling 712100, Shaanxi, People’s Republic of China

                [5 ]Author for correspondence: karin.tremetsberger@ 123456boku.ac.at
                Article
                apps1700081
                10.3732/apps.1700081
                5664967
                a91508c9-a80f-4acb-885f-a204f45a05c0
                © 2017 Wang et al. Published by the Botanical Society of America

                This is an open access article distributed under the terms of the Creative Commons Attribution License ( CC-BY-NC-SA 4.0), which permits unrestricted noncommercial use and redistribution provided that the original author and source are credited and the new work is distributed under the same license as the original.

                History
                : 29 July 2017
                : 25 August 2017
                Categories
                Primer Note

                asteraceae,clonal growth,hybridization,hypochaeris incana,perennial herb,polyploidy,south america

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