Nuclear DNA Content in Twelve Species of Alstroemeria L. and Some of their Hybrids

Nuclear DNA content (2C-value), estimated through flow cytometryusing propidium iodide (PI), was shown to vary from 36.5 pgto 78.9 pg among 29 accessions of 12Alstroemeria species (2n=2x =16). The extremes were found inA. magnifica ssp.magnificaand inA. ligtu ssp.simsii , both belonging to the Chilean speciesgroup. The four Brazilian species exhibited less variation innuclear DNA content (49.8–56.4 pg), than the eight Chileanspecies (36.5–78.9 pg). Nuclear DNA content was positivelycorrelated (r =0.92,n =7,P <0.01) with the total chromosomelength. It was also positively correlated (r =0.85,n =5,P <0.01)with the length of C-bands, when only the Chilean species wereconsidered. When both karyotype parameters, length of non-C-bandedchromosome regions (x) and length of C-bands (y) were determined,it was possible to predict the nuclear DNA content (z) withthe formula z=0.65x +1.31y-0.45 (R ²=0.97,P =0.004). The DAPI fluorescence of most accessions was proportional tothe PI fluorescence (r =0.98,P <0.001), except for one accessionofA. ligtu , that had a relatively high PI/DAPI ratio (1.88).The PI/DAPI ratios of the Brazilian species were lower (1.59–1.67)than those of the Chilean species (1.68–1.88), which mightreflect a difference in base pair composition. Four groups ofspecies could be distinguished on the basis of fluorescencevalues. Diploid interspecific hybrids were shown to have a DNAcontent intermediate to the values of the parents involved.Both the PI and the DAPI fluorescence values of these hybridsapproximated the mid parent values. Tetraploids, derived fromselfing of diploids, had PI and DAPI fluorescence values thatwere twice that of the diploid hybrids. It was possible to distinguishaneuploids from euploids based on fluorescence values. Alstroemeria ; aneuploidy; C-banding; DAPI; evolution; flow cytometry; genome size; geophytes; karyotypes; Inca Lily; nuclear DNA; propidium iodide     

Characterisation of distant Alstrogmeria hybrids: application of highly repetitive DNA sequences from A. ligtu ssp. ligtu

Clones from a Sau 3A family of eight highly repetitive sequences previously isolated from a genomic DNA library of Alstroemeria ligtu ssp. ligtu were sequenced and found to be highly conserved. A trinucleotide microsatellite repeat [GCA]_3?4 was present. A second, unrelated, Sau 3A repeat was also characterised. Southern analysis proved that the isolated repeats were specific for the A. ligtu subspecies and could not be detected in other Chilean or Brazilean Alstroemeria species. As shown by in situ hybridisation, the Sau 3A family and the unrelated Sau 3A repeat co‐localised at distinct sites along most chromosomes of Alstroemeria ligtu ssp. ligtu and Alstroemeria ligtu ssp. simsii. The present set of species‐specific repetitive sequences enables the identification of A. ligtu chromosomes, and thus the tracking of chromosome transmission to interspecific hybrids and their progeny.     

Genome size variation and C-band polymorphism inAlstroemeria aurea, A. ligtu andA. magnifica (Alstroemeriaceae)

Among a total of 43 accessions ofAlstroemeria aurea, A. ligtu andA. magnifica nuclear DNA amounts (2C-values) showed significant intraspecific variation, 1.09, 1.21 and 1.15 fold, respectively, when determined through flow cytometric measurements of fluorescence of propidium iodide (PI) stained nuclei. After staining with another fluorochrome, 4,6-diamidino-2-phenylindole (DAPI), an intraspecific variation of 1.10, 1.11 and 1.12 fold, respectively, was found. C-band polymorphisms were present among and within the accessions of all three species. In some cases only very small differences in C-banding pattern were observed. In other cases, however, differences were more prominent. Besides C-band polymorphism, there were also instances of chromosome length polymorphism, which concerned the total chromosome complement or single chromosomes. The variation in nuclear DNA amount inA. aurea andA. ligtu was more or less continuous, except for one accession ofA. ligtu subsp.simsii. Artificial selection and possibly introgression of chromosomes from other species may have moulded the karyotypes of some of the accessions ofA. aurea, a species that has been under cultivation for more than 160 years. The variation as observed inA. magnifica subsp.magnifica was discontinuous and could be due to a broad species concept.     

Constitutive heterochromatin DNA polymorphisms in diploid Medicago sativa ssp. falcata.

A Giemsa C-banding technique was used to study the amount and location of constitutive heterochromatin in diploid (2n = 2x = 16) Medicago sativa ssp. falcata (L.) Arcangeli. Most accessions had the standard C-banding pattern with centromeric bands on all the chromosomes and a prominent heterochromatic band at the nucleolar organizer regions (NOR) of the satellited (SAT) chromosomes. However, we observed in various accessions that constitutive heterochromatic C-bands can exist at the telomeric ends of all the chromosomes. Interstitial bands occurred on the short arms of all chromosomes except for chromosome 3 and on the long arms of chromosomes 1, 2, 3, and 6, only. Rearranged chromosomes such as isochromosomes have been observed for the short arms of chromosomes 2 and 6. This is the first report on the existence of C-banding polymorphisms and the detection of putative isochromsomes in the chromosomes of diploid ssp. falcata which could have contributed to the variation observed in cultivated alfalfa.     

Giemsa C-banding patterns in Aedes (Stegomyia) mosquitoes

Chromosomes from gonads of 14–24 h old pupae of nine species of Stegomyia mosquitoes have been examined using the Giemsa C-banding technique. The species studied were Aedes albopictus, A. polynesienis, A. scutellaris, A. alcasidi, A. seatoi, A. pseudalbopictus, A. melallicus, A. annandalei and A. vittatus. The diploid chromosome number of all species is six. All species possess C-bands in the centromeric regions of each of the three pairs of chromosomes. Besides, an intercalary C-band is present on the female determining (=m) chromosome but absent from the male—determining (= M) chromosome of all species except A. vittatus. In A. vittatus, the m and M chromosomes possess a terminal C-band. Thus, the nine species of Aedes analysed showed two distinct patterns of C-banding. —The evolution of heterochromatin patterns in various species is also discussed. The Giemsa C-banding technique should prove useful in studies of chromosomal speciation in culicine mosquitoes.     

Satellite DNA probes of Alstroemeria longistaminea (Alstroemeriaceae) paint the heterochromatin and the B chromosome,reveal a G-like banding pattern,and point to a strong structural karyotype conservation

Alstroemeria species present a well-conserved and asymmetric karyotype. The genus is divided into a Chilean clade, rich in heterochromatin, and a Brazilian clade, poor in heterochromatin. We investigated the distribution of the main repetitive sequences in the chromosomes of the Brazilian species A. longistaminea (2n = 16 + 0-6B) aiming to evaluate the role played by these sequences on the structural organization of the karyotype. In situ hybridization of the three most abundant retrotransposons, corresponding to ~ 45% of the genome, was uniformly distributed. Three satellite DNA sequences, representing near half of the whole satellite fraction (1.93% of the genome), were mainly concentrated on the heterochromatin and one of them painted the whole B chromosome. Noteworthy, some satellites were located on euchromatin, either dispersed or concentrated in clusters along the chromosomes, revealing a G-band-like pattern. The two satellites that presented more C-band- and G-band-like labeling were also hybridized in situ in two other Alstroemeria species. They revealed astonishing similar patterns of distribution, indicating an unusually structural karyotype conservation among Brazilian species.

     

C-banding analysis of eight species ofKengyilia (Poaceae: Triticeae)

To characterize chromosomes and the interspecific relationships within the genus Kengyilia, 8 species were used for Giemsa C-banding analysis. Results indicated that the species differed in C-banding patterns. K. gobicola, K. alatavica and K. batalinii had distinct centromeric bands and no banded chromosomes, while K. hirsuta, K. longiglumis, K. melanthera, K. rigidula and K. thoroldiana had more abundant and diagnostic C-bands with interstitial and terminal bands.     

Chromosome Banding Patterns in Some Indian Pulses

By using the Giemsa C-banding technique, chromosome bandingpatterns on the somatic chromosomes of eight important pulsecrops, pea, lentil, guar (cluster bean), chick pea, pigeon pea,mung bean (green gram), urd (black gram) and cowpea have beenstudied. Each species has a characteristic C-banding pattern.The significance of such banding patterns which correlate withthe position of pachytene knobs, in chromosome identification,and in assigning relationships at the cytological level in thepulses of genus Vigna is stressed. Chromosome banding, Giemsa C-banding, pulse crops, Pisum sativum L., garden pea, Lens culinaris Medik, lentil, Cyamopsis tetragonoloba (L.) Taub., guar, Cicer arietinum L., chick pea, gram, Cajanus cajan (L.) Millsp., pigeon pea, Vigna radiata (L.) Wilczek, mung bean, Vigna mungo (L.) Hepper, urd, Vigna unguiculata (L.) Walp, cowpea     

Giemsa C-banded karyotypes and taxonomic relationships of some North AmericanAllium species and their relationship to Old World species (Liliaceae)

The somatic karyotypes of six North AmericanAllium species and the EuropeanA. scorzonerifolium have been investigated using a Giemsa C-banding technique. All species have a chromosome number of 2n = 14. InA. scorzonerifolium and the three North American speciesA. dichlamydeum, A. fibrillum andA. unifolium C-bands are restricted to two pairs of nucleolar chromosomes. Each chromosome has a band proximal to the nucleolar constriction and a positively banded satellite. InA. acuminatum, in addition to the bands associated with the nucleolar constrictions, all chromosomes also have pericentromeric bands.A. cernuum exhibits a distinctive banding style: two chromosome pairs with bands adjacent to the nucleolar constrictions and four pairs with telomeric bands on their short arms. In the karyotype ofA. geyeri neither C-bands nor nucleolar chromosomes were found.—A comparison of the banding styles together with other cytological and morphological characters of these species with old world members ofAllium reveals:A. cernuum closely resembles species within subgenusRhizirideum, whereas the other species studies exhibit many similarities with subgenusMolium. Their sectional grouping and their relationships with Old World species are discussed.     

茅舍血厉螨核型及染色体的C带、G带的研究

本文首次报道了一种革螨——茅舍血厉螨核型及染色体C带、G带的研究。用剖腹取卵法、玻璃纸压片、Giemsa染色,经分析茅舍血厉螨的核型,单倍体n=7,二倍体2n=14。 用氢氧化钡—吉姆萨技术显示茅舍血厉螨染色体C带。在第1、2、4、5染色体上出现恒定的C带部分,第3、6、7染色体上出现不恒定的C带部分。根据C带带型,茅舍血厉螨着丝点的位置可分为近中区域(sm),近端区域(St),末端区域(t)和末端点(T)四类。 G带分析用胰蛋白酶—吉姆萨技术显示。 本文对茅舍血厉螨的核型、革螨染色体研究中螨卵的收集方法和染液的改进、C带带型与着丝粒位置的确定和G带显带问题进行了讨论。     

Characterization of different types of condensed chromatin inCostus (Zingiberaceae)

The chromatin structure of six diploids species ofCostus was analysed using conventional Giemsa staining, C-banding and DAPI/CMA fluorochromes. The interphase nuclei in all the species show an areticulate structure and the prophase chromosomes show large blocks of proximal condensed chromatin. After banding procedures, each chromosome exhibits only centromeric dot-like DAPI⁺/CMA^– C-bands whereas the satellites (one pair at each karyotype) are weakly stained after C-banding and show a DAPI^–/CMA⁺ fluorescence. Two chromocentres show bright fluorescence with CMA and weak staining after C-banding whereas the others chromocentres show only a small fraction of DAPI⁺ heterochromatin. These results were interpreted to mean that the greater part of the condensed chromatin has an euchromatic nature whereas two types of well localized heterochromatin occur in a small proportion. The Z-stage analysis suggests that heterochromatin and condensed euchromatin decondense at different times. The chromosome number and morphology of all species are given and the implications of the condensed euchromatin are discussed.Dedicated to Prof.Elisabeth Tschermak-Woess on the occasion of her 70th birthday.     

玉米8个栽培亚种（类型）的核型和C—带带型的比较研究

本文首次报道了玉米8个亚种、2个亚型和2个杂交品种的核型和Giemsa C-带带型。所有材料的根尖细胞染色体数目均为2n=20。主要由中部和亚中部着丝点染色体组成。第6染色体短臂均具随体,但大小不同。所有材料均显示有亚端带和端带,在第6染色体的短臂上显示有NOR或/和随体带。C-带的分布、总数目和总长度各不相同。其总带数变异于6至18之间,C-带总长度为5.65—11.40％之间。在核型中,具中部着丝点的染色体数目及C-带总数,罕见栽培或原始的类型通常多于广泛栽培的类型。此外,有关核型和C-帝的变异和进化也进行了简略的讨论。     

Cytogenetics of bisexual/unisexual species of Poecilia. I. C-bands, Ag-NOR polymorphisms, and sex chromosomes in three populations of Poecilia latipinna

Three populations of the North American cyprinodont fish Poecilia latipinna, considered to be one of the progenitor species of the gynogenetic unisexual P. formosa, were analyzed by C-banding and Ag-staining. C-bands were found to be polymorphic, and Ag-staining showed a high degree of variability in both the number and location of nucleolus organizer regions. The C-banding and Ag-staining patterns allow, to a certain extent, to distinguish individual specimens from each of these populations. Females of the three populations were found to have a heteromorphic chromosome pair, which was frequently identifiable with Giemsa staining and always after C-banding. This pair could be interpreted as sex chromosomes of the ZW/ZZ type.     

Cytochemical evaluation of C-heterochromatic-DNA in metaphase chromosomes.

A method is proposed to evaluate the amount of DNA resistant to the C-banding pretreatments (C-heterochromatic-DNA) in metaphase chromosomes. Measurements were performed by microfluorometry on propidium iodide stained metaphases of man, gorilla and mouse; in these species, C-heterochromatin exhibits significant differences of both base composition and distribution along the chromosomes. The amount of C-heterochromatic-DNA was found to be about 16%, 28% and 58% of the total DNA content (genome size) in man, gorilla and mouse, respectively. The areas of C-bands after Giemsa staining were also assessed by microdensitometry, and corresponded to about 8%, 15% and 14% of the total karyotype area of man, gorilla and mouse respectively. No direct relation thus exists between C-band areas and the amount of DNA resistant to the C-banding pretreatments. In man and gorilla, the amount of C-heterochromatic-DNA accounts for the differences observed in genome size.     

Restriction endonuclease banding of rainbow trout chromosomes

Rainbow trout chromosomes were treated with nine restriction endonucleases, stained with Giemsa, and examined for banding patterns. The enzymes AluI, MboI, HaeIII, HinfI (recognizing four base sequences), and PvuII (recognizing a six base sequence) revealed banding patterns similar to the C-bands produced by treatment with barium hydroxide. The PvuII recognition sequence contains an internal sequence of 4 bp identical to the recognition sequence of AluI. Both enzymes produced centromeric and telomeric banding patterns but the interstitial regions stained less intensely after AluI treatment. After digestion with AluI, silver grains were distributed on chromosomes labeled with [³H]thymidine in a pattern like that seen after AluI-digested chromosomes are stained with Giemsa. Similarly, acridine orange (a dye specific for DNA) stained chromosomes digested with AluI or PvuII in patterns resembling those produced with Giemsa stain. These results support the theory that restriction endonucleases produce bands by cutting the DNA at specific base pairs and the subsequent removal of the fragments results in diminished staining by Giemsa. This technique is simple, reproducible, and in rainbow trout produces a more distinct pattern than that obtained with conventional C-banding methods.     

Comparison of the Giemsa C-Band karyotypes and the relationships of Allium cepa,A. fistulosum and A. galanthum

Giemsa C-band staining of somatic chromosomes in Allium cepa, A. cepa cv. Shallot, A. fistulosum and A. galanthum shows that these four taxa are characterised by all chromosomes having one band at each telomere, except chromosome arms with satellites, in which case the band is proximal to the nucleolar constriction. This similarity is correlated with common morphological characteristics and an ability of the three species to hybridize with one another, indicating that they have a close genetic relationship. The results indicate that Giemsa C-bands are evolutionarily conservative in this group and provide a useful criterion for determining relationships.     

杉木种子中的两种C带型

在裸子植物中,与随体有关的性染色体的研究已有一些报道。其中陈瑞阳提到,在银杏中,用 Giemsa C 带方法揭示,雌性是异型的,性染色体对的其中一个比另一个具有明显大的随体和核仁缢痕带。而雄性是同型的,性染色体对具有同等大小的随体和核仁缢痕带。     

Chromosome homology and heterochromatin in goat,sheep and ox studied by banding techniques

Peripheral blood lymphocyte metaphase chromosomes of three Bovoidean species have been studied using Quinacrine fluorescence and Giemsa banding techniques to give Q-, G-, and C-banding patterns. Q- and G-banding characteristics, coupled with chromosome length, enabled all of the chromosomes in each of the chromosome complements to be clearly distinguished, although some difficulties were encountered with the very smallest chromosomes. A comparison of G-banding patterns between the species revealed a remarkable degree of homology of banding patterns. Each of the 23 different acrocentric autosomes of the domestic sheep (2n=54) was represented by an identical chromosome in the goat (2n=60) and the arms of the 3 pairs of sheep metacentric autosomes were identical matches with the remaining 6 goat acrocentrics. A similar interspecies homology was evident for all but two of the autosomes in the ox (2n=60). This homology between sheep metacentric and goat acrocentric elements confirms a previously suggested Robertsonian variation. The close homology in G-banding patterns between these related species indicates that the banding patterns are evolutionarily conservative and may be a useful guide in assessing interspecific relationships. —The centromeric heterochromatin in the autosomes of the three species was found to show little or no Q-or G-staining, in contrast to the sex chromosomes. This lack of centromeric staining with the G-technique (ASG) contrasts markedly with results obtained with other mammalian species. However, with the C-banding technique these regions show a normal intense Giemsa stain and the C-bands in the sex chromosomes are inconspicuous. The amount of centromeric heterochromatin in the sheep metacentric chromosomes is considerable less than in the acrocentric autosomes or in a newly derived metacentric element discovered in a goat. It is suggested that the pale G-staining of the centromeric heterochromatin in these species might be related to the presence of G-Crich satellite DNA.     

黄颡鱼染色体的C─带、Ag—NORs、荧光带和复制带显带的研究

采用Giemas染色、C─带、Ag—NORs、荧光染色和复制带显带的技术对黄颡鱼染色体进行了研究。结果表明,黄颡鱼只有部分的染色体呈现阳性C─带,可分为三类,其中NORs区是染色最深、染色面积最大的区域,为深染居间C─带。其Ag－NORs位于m5q末端。CMA3染色显示NORs区呈现出明亮的荧光。中复制染色体上着丝粒区、端粒区和居间区浅染。发现核仁缢痕、深染居间C─带、Ag—NORs、CMA3明亮区和中复制带浅染NORs区位置基本一致,C─带阳性区和中复制带浅染区具有对应性。