A data base for PCR primers in the chloroplast genome
The data base, and some applications

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Introduction
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The data base, and some applications
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The data base, and some applications

Currently, the complete database contains ~ 500 primers, mainly in the 'long single copy' (LSC) region. In our hands, many of the published and newly designed primers can be used in different combinations, and in many species. 'Generic' PCR conditions that favour sucessful amplification for majority of combinations are: 2 mM Mg2+; and a PCR program with 10 cycles @ 70 deg annealing, followed by 32 cycles @ 55 deg annealing. With these primers and methods, the chloroplast genome of hitherto unexplored speies can be scanned in detail, and regions specific for different purposes can be picked.

Detailed maps of the primer sequences on the tobacco chloroplast can be found following this link.

A table with 356 primer sequences can be found here.
Primer sequence data are presented in a simple tabular format. It is intended to install also a direct ordering service for visitors to this website (please contact me if you are from a company interested in participating in such a service). Please contact me if you have any further questions.

Some examples of the use of 'universal'' primers

Populus restriction maps around ycf9 (Heinze 1998):
In this genus, a region close to the ycf9 genes contained a number of species- or clone-specific polymorphisms, which could be clarified by simple restriction mapping (picture). On the basis of these results, simple tests for rapid species identification could be developed (Krystufek 2001).
Cherry restriction maps (Prunus avium L.; Heinze, unpublished)
In this species, many small indels occur. Some of the seem to be clustered in the trnR-atpA region. Restriction mapping of overlapping PCR fragments revealed location of the indels and allowed greater resolution for haplotypes.
Denaturing HPLC analysis of chloroplast DNA variation
Heteroduplex analysis was conducted with a VARIAN gradient HPLC system. Heteroduplices partially melt at critical temperature and give an indicative pattern in this system. Point mutations in fragments up to 700 bp can be detected. The system is especially suitable for screening large number of samples.
Mutation hotspots ?
Our preliminary data reveal that the junction of the inverted repeat A and the large single copy region is polymorphic within many species. Several authors have found polymorphisms in ths region (Jackson et al. 1999). Other regions are sometimes highly polymorphic, but only for specific taxa (e.g., the ycf9 region in Populus).

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