The pachytene chromosomes of maize as revealed by fluorescence in situ hybridization with repetitive DNA sequences

Abstract

A repetitive DNA sequence, ZmCR2.6c, was isolated from maize based on centromeric sequence CCS1 of the wild grass Brachypodium sylvaticum. ZmCR2.6c is 309 bp in length and shares 65% homology to bases 421-721 of the sorghum centromeric sequence pSau3A9. Fluorescence in situ hybridization (FISH) localized ZmCR2.6c to the primary constrictions of pachytene bivalents and to the stretched regions of MI/AI chromosomes, indicating that ZmCR2.6c is an important part of the centromere. Based on measurements of chromosome lengths and the positions of FISH signals of several cells, a pachytene karyotype was constructed for maize inbred line KYS. The karyotype agrees well with those derived from traditional analyses. Four classes of tandemly repeated sequences were mapped to the karyotype by FISH. Repeats 180 bp long are present in cytologically detectable knobs on 5L, 6S, 6L, 7L, and 9S, as well as at the termini and in the interstitial regions of many chromosomes not reported previously. A most interesting finding is the presence of 180-bp repeats in the NOR-secondary constriction. TR-1 elements co-exist with 180-bp repeats in the knob on 6S and form alone a small cluster in 4L. 26S and 5S rRNA genes are located in the NOR and at 2L.88, respectively. The combination of chromosome length, centromere position, and distribution of the tandem repeats allows all chromosomes to be identified unambiguously. The results presented form an important basis for using FISH for physical mapping and for investigating genome organization in maize.

High-Resolution Single-Copy Gene Fluorescence in Situ Hybridization and Its Use in the Construction of a Cytogenetic Map of Maize Chromosome 9

“…This cocktail is composed of centromeric repeat (CentC) (Ananiev et al, 1998b), telomere sequences, 5S and 45S rRNA genes, and the 180-bp (Peacock et al, 1981) and TR-1 (Ananiev et al, 1998a) knob repeated sequences. These repetitive sequences have been localized on maize pachytene chromosomes in separate experiments (Chen et al, 2000;Hsu et al, 2003;Wang and Chen, 2003). To place all these markers on chromosomes in a single FISH experiment, six different repetitive sequences were labeled with either one or two fluorochromes and were hybridized to pachytene chromosomes of the Kansas Yellow Saline (KYS) inbred line.…”

“…Meiotic pachytene chromosomes have been shown to be a better target for high-resolution FISH mapping in plants (Xu and Earle, 1996;Zhong et al, 1996aZhong et al, , 1996bFransz et al, 1998;Chen et al, 2000). First, pachytene chromosomes are generally 10 to 20 times longer than somatic metaphase chromosomes and possess many cytological landmarks Chen et al, 2000;see Supplemental Table 1 online).…”

“…First, pachytene chromosomes are generally 10 to 20 times longer than somatic metaphase chromosomes and possess many cytological landmarks Chen et al, 2000;see Supplemental Table 1 online). Second, the wall of pollen mother cells can be easily digested by enzyme treatment, thus facilitating probe penetration.…”

“…Chen et al (2000) demonstrated that a mild squash procedure produced high-quality pachytene chromosomes suitable for high-resolution mapping with repetitive sequence probes. Here, we describe a set of probes for unambiguous identification of the 10 maize pachytene chromosomes and compare the karyotypes derived from the squash and 3-D chromosome preparations to evaluate the preservation of chromosome morphology after squash procedure.…”

Estudios citogenéticos evolutivos del género Zea

“…A nivel de secuencias, en la región de los knobs se encontraron dos familias de DNA repetidas en tandem, una de 180 pares de bases y otra de 350 pares de bases, también denominada TR-1 (Chen et al, 2000).…”