Towards a Transferable and Cost-Effective Plant AFLP Protocol

Amplified fragment length polymorphism (AFLP) is a powerful fingerprinting technique that is widely applied in ecological and population genetic studies. However, its routine use has been limited by high costs associated with the optimization of fluorescently labelled markers, especially for individual study systems. Here we develop a low-cost AFLP protocol that can be easily transferred between distantly related plant taxa. Three fluorescently labelled EcoRI-primers with anchors that target interspecifically conserved genomic regions were used in combination with a single non-labelled primer in our AFLP protocol. The protocol was used to genotype one gymnosperm, two monocot and three eudicot plant genera representing four invasive and four native angiosperm species (Pinus pinaster (Pinaceae), Pennisetum setaceum and Poa annua (Poaceae), Lantana camara (Verbenaceae), Bassia diffusa (Chenopodiaceae), Salvia lanceolata, Salvia africana-lutea, and Salvia africana-caerulea (Lamiaceae)). Highly polymorphic and reproducible genotypic fingerprints (between 37–144 polymorphic loci per species tested) were obtained for all taxa tested. Our single protocol was easily transferred between distantly related taxa. Measures of expected heterozygosity ranged from 0.139 to 0.196 for P. annua and from 0.168 to 0.272 for L. camara which compared well with previously published reports. In addition to ease of transferability of a single AFLP protocol, our protocol reduces costs associated with commercial kits by almost half. The use of highly conserved but abundant anchor sequences reduces the need for laborious screening for usable primers that result in polymorphic fingerprints, and appears to be the main reason for ease of transferability of our protocol between distantly related taxa.

[1]  D. Ingram,et al.  Plant tissue culture , 1976 .

[2]  E. Pahlich,et al.  A rapid DNA isolation procedure for small quantities of fresh leaf tissue , 1980 .

[3]  C. Hutchinson,et al.  Poa Annua L. , 1982 .

[4]  J. Doyle,et al.  A rapid DNA isolation procedure for small amounts of fresh leaf tissue , 1987 .

[5]  M. Lynch,et al.  Analysis of population genetic structure with RAPD markers , 1994, Molecular ecology.

[6]  P. Vos,et al.  AFLP: a new technique for DNA fingerprinting. , 1995, Nucleic acids research.

[7]  J. Hamrick,et al.  Effects of life history traits on genetic diversity in plant species , 1996 .

[8]  R. Van der Hoeven,et al.  Visualization of differential gene expression using a novel method of RNA fingerprinting based on AFLP: analysis of gene expression during potato tuber development. , 1996, The Plant journal : for cell and molecular biology.

[9]  J. Ruiz-Herrera,et al.  Differences in DNA methylation patterns are detectable during the dimorphic transition of fungi by amplification of restriction polymorphisms , 1997, Molecular and General Genetics MGG.

[10]  P. VanRaden,et al.  Standardization and conversion of marker polymorphism measures. , 1999, Animal biotechnology.

[11]  L. Xiong,et al.  Patterns of cytosine methylation in an elite rice hybrid and its parental lines, detected by a methylation-sensitive amplification polymorphism technique , 1999, Molecular and General Genetics MGG.

[12]  U. Mueller,et al.  AFLP genotyping and fingerprinting. , 1999, Trends in ecology & evolution.

[13]  A. Melchinger,et al.  Two high-density AFLP® linkage maps of Zea mays L.: analysis of distribution of AFLP markers , 1999, Theoretical and Applied Genetics.

[14]  G. Mueller-Warrant,et al.  Genetic diversity of Poa annua in western Oregon grass seed crops , 2000, Theoretical and Applied Genetics.

[15]  M. De Loose,et al.  AFLP based alternatives for the assessment of Distinctness, Uniformity and Stability of sugar beet varieties , 2001, Theoretical and Applied Genetics.

[16]  D. Chagné,et al.  Genetic diversity within and among Pinus pinaster populations: comparison between AFLP and microsatellite markers , 2001, Heredity.

[17]  C. G. van der Linden,et al.  Biodiversity assessment using markers for ecologically important traits , 2002 .

[18]  I. Roldán‐Ruiz,et al.  Data from amplified fragment length polymorphism (AFLP) markers show indication of size homoplasy and of a relationship between degree of homoplasy and fragment size , 2002, Molecular ecology.

[19]  Lang Shen,et al.  Genetic structure along a gaseous organic pollution gradient: a case study with Poa annua L. , 2003, Environmental pollution.

[20]  P. Taberlet,et al.  Using AFLP to resolve phylogenetic relationships in a morphologically diversified plant species complex when nuclear and chloroplast sequences fail to reveal variability. , 2003, Molecular phylogenetics and evolution.

[21]  Christian Schlötterer,et al.  The evolution of molecular markers — just a matter of fashion? , 2004, Nature Reviews Genetics.

[22]  Method for Quality DNA Isolation from Different Parts of Jute Plant: Corchorus capsularis L. and C. olitorius L. , 2004 .

[23]  P. Taberlet,et al.  How to track and assess genotyping errors in population genetics studies , 2004, Molecular ecology.

[24]  P. Taberlet,et al.  Genotyping errors: causes, consequences and solutions , 2005, Nature Reviews Genetics.

[25]  Z Zhang,et al.  Analysis of allele fidelity, polymorphic information content, and density of microsatellites in a genome-wide screening for hip dysplasia in a crossbreed pedigree. , 2005, The Journal of heredity.

[26]  S. Weller,et al.  Genetic diversity does not affect the invasiveness of fountain grass (Pennisetum setaceum) in Arizona, California and Hawaii , 2005 .

[27]  François Pompanon,et al.  Use of amplified fragment length polymorphism (AFLP) markers in surveys of vertebrate diversity. , 2005, Methods in enzymology.

[28]  S. Bensch,et al.  Ten years of AFLP in ecology and evolution: why so few animals? , 2005, Molecular ecology.

[29]  M. Fay,et al.  The effects of nuclear DNA content (C-value) on the quality and utility of AFLP fingerprints. , 2005, Annals of botany.

[30]  R. Harcourt,et al.  Widespread utility of highly informative AFLP molecular markers across divergent shark species. , 2006, The Journal of heredity.

[31]  P. Smouse,et al.  genalex 6: genetic analysis in Excel. Population genetic software for teaching and research , 2006 .

[32]  J. L. Le Roux,et al.  Super-Genotype: Global Monoclonality Defies the Odds of Nature , 2007, PloS one.

[33]  S. Manel,et al.  Statistical analysis of amplified fragment length polymorphism data: a toolbox for molecular ecologists and evolutionists , 2007, Molecular ecology.

[34]  Heidi M. Meudt,et al.  Almost forgotten or latest practice? AFLP applications, analyses and advances. , 2007, Trends in plant science.

[35]  K. Chwedorzewska.  Poa annua L. in Antarctic: searching for the source of introduction , 2008, Polar Biology.

[36]  M. V. Eijk,et al.  AFLP technology for DNA fingerprinting , 2007, Nature Protocols.

[37]  Katrina M. Dlugosch,et al.  Molecular and quantitative trait variation across the native range of the invasive species Hypericum canariense: evidence for ancient patterns of colonization via pre‐adaptation? , 2007, Molecular ecology.

[38]  Jinfa F. Zhang,et al.  ATG-anchored AFLP (ATG-AFLP) analysis in cotton , 2008, Plant Cell Reports.

[39]  A. Wieczorek,et al.  Molecular systematics and population genetics of biological invasions: towards a better understanding of invasive species management , 2009 .

[40]  T. Mahmood,et al.  AFLP mediated genetic diversity of malvaceae species , 2010 .

[41]  E. Bornberg-Bauer,et al.  Evolutionary divergence and limits of conserved non-coding sequence detection in plant genomes , 2011, Nucleic acids research.

[42]  Cost-effective fluorescent amplified fragment length polymorphism (AFLP) analyses using a three primer system , 2011 .

[43]  Isolation and characterization of polymorphic microsatellite loci from the invasive plant Lantana camara L. , 2012, Conservation Genetics Resources.

[44]  Heidi M. Meudt,et al.  Cost‐effective fluorescent amplified fragment length polymorphism (AFLP) analyses using a three primer system , 2011, Molecular ecology resources.

[45]  D. Richardson,et al.  Management history determines gene flow in a prominent invader , 2013 .

[46]  R Core Team,et al.  R: A language and environment for statistical computing. , 2014 .