Popgene Version 1.31

Applications in Plant Sciences Published by: Botanical Society of America. (H e) were calculated using the software POPGENE version 1.31 (Yeh et al.,1999). Characteristics of 11 polymorphic microsatellite primers developed for Lagerstroemia indica ‘Hong Die Fei Wu’. How can I determine the genetic diversity of a wild plant population? POPGENE version 1.31, use this software and GenAlex. POPGENE Software for Population Genetic Analysis.

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Since the 1960s, L. Fauriei Koehne has played an important role in crape myrtle breeding programs because of its strong resistance to mildew diseases and to cold temperatures. Lagerstroemia speciosa (L.) Pers., L. Limii Merr., and L. Subcostata Koehne have also been introduced into crape myrtle breeding programs recently (; ). Undoubtedly, many Lagerstroemia cultivars and species with excellent ornamental traits will bring great changes to crape myrtle breeding. Therefore, to improve the usefulness of molecular marker-assisted selection programs in Lagerstroemia, suitable molecular markers are needed to identify, assess, conserve, and use these germplasms of Lagerstroemia.

For such an objective, simple sequence repeats (SSRs) have proven to be effective and useful for the evaluation of genetic diversity among Lagerstroemia species and cultivars (;; ) because of their codominance and hypervariablity. However, available SSR markers are relatively limited in crape myrtle (;; ). Here we report the rapid development of 11 SSR markers and their cross-species transferability. METHODS AND RESULTS Samples of L. Indica cultivars and related species were cultivated in the crape myrtle collection of the China National Engineering Research Center for Floriculture, Beijing (40°02′13.67″N, 115°50′5.58″E) ().

Genomic DNA was extracted from silica-dried leaf tissue with the DNAsecure Plant Kit following the manufacturer's protocol (Tiangen Biotech, Beijing, China). A microsatellite-enriched library was constructed following a modified biotin-streptavidin capture method (). In brief, the genomic DNA of L.

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Indica ‘Hong Die Fei Wu’ was digested into segments with the restriction enzymes RsaI and XmnI (New England Biolabs, Beijing, China), ligated to the double-stranded Super SNX-24 linker (F: 5′-GTTTAAGGCCTAGCTAGCAGAATC-3′, R: 5′-pGATTCTGCTAGCTAGGCCTTAAACAAA-3′; synthesized by Sangon Biotech, Shanghai, China), amplified using PCR, then hybridized with a mix of 3′ biotin-labeled oligonucleotide probes and captured for microsatellites using streptavidin-coated magnetic beads (Dynabeads M-280; Invitrogen, Carlsbad, California, USA). The captured DNA was amplified by PCR reaction using the Super SNX-24 forward linker as primer. The enriched DNA was inserted into pCR2.1-TOPO vectors (Invitrogen) following the manufacturer's instructions and transformed into One Shot Top10 Chemically Competent cells (Invitrogen).

Recombinant clones were identified using blue/white screening on Luria-Bertani agar plates containing ampicillin and Xgal. A total of 175 bacterial colonies were picked out and analyzed using M13 primers to amplify the complete microsatellite-containing insert. Secret discovery rapidshare. SSR-containing clones were selected as positives when one well band was visible on a 2% agarose gel after PCR. Then, the positive clones were sequenced on an ABI 3730 DNA analyzer (Applied Biotech, San Diego, California, USA). Finally, sequence analysis was carried out using the EditSeq of the DNASTAR software package (DNASTAR, Madison, Wisconsin, USA). SSR loci were located using the program SSRHunter 1.3.0 (Qiang Li, Nanjing Agricultural University, Nanjing, China).