|A data base for PCR primers in the chloroplast genome|
A Data Base of PCR Primers for the Study of the Chloroplast Genome in Plants
Berthold Heinze, Federal Research Centre for Forests - Department of Genetics, Vienna, AUSTRIAVersion 2.1 (March 2007 - more than 700 primer sequences)
- Heinze B., 2007: A database of PCR primers for the chloroplast genomes of higher plants. Plant Methods 3: 4.
- Heinze B. , 2005: The chloroplast PCR primer database: tools for comprehensive phylogeographic analysis of a whole genome. Poster presented at International Botany Congress, 17-23 July 2005, Vienna, Austria/Europe. (follow the link to open the PDF of the poster!)
- Grivet, D., Heinze, B., Vendramin, G. G., and Petit, R. J. Genome walking with consensus primers: application to the large single copy region of chloroplast DNA. Molecular Ecology Notes 1(4), 345-349. 2001.
- Heinze B. , 2001:
A data base
for PCR primers in the chloroplast genome. Poster presented at Plant and
Animal Genome X, Jan 12-16, 2002, San Diego, CA, USA.
- Heinze, B.; Grivet, D.; Petit, R.; Hahn, B , 1999: A comprehensive set of PCR primers for the study of the chloroplast genome in plants. "Networking Day Genomics" (BIT and BMBWK), Wien, Mai 1999. Poster
- Heinze, B. , 1999: Comparative chloroplast DNA investigations in some hardwood forest trees using PCR-RFLP analysis. Plant and Animal Genome VII, Jan. 17-21, , San Diego, USA , poster
IntroductionSeveral authors have described the use of "universal" primers for PCR from chloroplast DNA, i.e., primers that amplify DNA fragments from many species in a genus, family, or even higher taxonomic groupings (see dedicated list of references). Usually, such primers are based on conserved DNA sequence in exons. The information is contained in a few comprehensive articles and many scattered primer sequences in other publications.
For use in molecular population genetics and taxonomy, large numbers of markers are necessary. Their (known) levels of polymorphism should fit diverse applications. They should be applicable for many species, and lend temselves to rapid screening methods.
However, marker development for such 'universal PCR' is time-consuming, so markers that can be transferred between species are desirable. This is possible with 'classical' RFLP probes. As shown by several authors, it is also possible with PCR primers, but to what extent ? The database compiled here will provide a starting ground to estimate primer 'universality'. In chloroplast spacers, gene order is highly conserved, and spacers show intra-species variation. Small indels predominate. Exon sequences are generally highly conserved, but this depends on the gene in question.
Some problems with universal PCR include the more rapid sequence evolution in some parts of the chloroplast; the identification of polymorphisms between conserved primer sites (introns); and occasional rearrangements, deletions, duplications in some genera, families, or higher taxonomic groups.
Sets of dedicated 'universal' chloroplast primers for amplifying spacers and introns have been described by Taberlet et al. 1991, Demesure et al. 1995, Dumolin-Lapegue et al. 1997, Fofana et al. 1997, Hamilton 1999, and Grivet et al. 2001. In the latter publication, the large single copy region of the chloroplast ring is covered by 38 primer pairs. Amplified fragments can be analysed by restriction or sequencing.
'Universal' chloroplast primers have also been developed for comparative sequencing in phylogeny, e.g. in the rbcL gene, for obtaining phylogenies of land plants. Other genes with different mutation rates can resolve certain problems, e.g. matK, ndhF, rpl16, and atpB.