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Construction of an Amaranthus hypochondriacus Bacterial Artificial Chromosome Library and Genomic Sequencing of Herbicide Target Genes

^a Dep. of Plant and Wildlife Sciences, Brigham Young Univ., Provo, UT, 84602
^b Arizona Genomics Institute, Dep. of Plant Sciences, Univ. of Arizona, Tucson, AZ 85721

^* Corresponding author (Jeff_Maughan{at}byu.edu).

Before the Spanish conquest of the ancient Americas, the grain amaranths (Amaranthus caudatus L., A. cruentus L., A. hypochondriacus L.) were a staple food of the New World. Recently, the grain amaranths have regained international attention for their nutritional quality and importance as a symbol of indigenous cultures. Here we report the development of a bacterial artificial chromosome (BAC) library constructed from the cultivar ‘Plainsman’ (A. hypochondriacus; 2n = 2x = 32). The library consists of a total of 36,864 clones with an average insert size of 147 kb with less than 1.8% of the clones containing empty vectors. The frequency of BAC clones carrying inserts derived from chloroplast and mitochondrial DNA was estimated to be 6.9%. Thus, based on a haploid genome size of 466 Mb per haploid nucleus, the BAC library coverage is approximately 10.6 times the haploid genome content. The genome coverage estimate was empirically confirmed by screening the library with seven low copy amaranth probe sequences. The utility of the amaranth BAC library was demonstrated by identification and full-length genomic sequencing of the acetolactate synthase and protoporphyrinogen oxidase genes—both major targets for several classes of important herbicides. The quality of the BAC library for BAC end sequencing projects was evaluated by bidirectional end sequencing of 384 random clones. End sequences were annotated using BLAST searches and queries to plant transposable element databases.

Abbreviations: ALS, acetolactate synthase • BAC, bacterial artificial chromosome • BES, BAC end sequence • CHEF, contour-clamped homogeneous electric field • HMW, high molecular weight • PCR, polymerase chain reaction • PPO, protoporphyrinogen oxidase • SSR, simple sequence repeat • UTR, untranslated region

Received for publication August 8, 2007.