Streptomycin and lincomycin resistances are selective plastid markers in cultured Nicotiana cells

Summary Resistance to streptomycin and lincomycin in plant cell culture is used as a color marker: resistant cells are green whereas sensitive cells are white on the selective medium. Streptomycin and lincomycin at appropriate concentrations do not kill sensitive Nicotiana cells. The selective value of plastid ribosomal DNA mutations, conferring resistance to streptomycin and lincomycin, was investigated by growing heteroplastidic cells on a selective medium. The heteroplastidic cells were obtained by protoplast fusion, and contained a mixed population of streptomycin resistant plastids from the N. tabacum line Nt-SR1-Kan2, and lincomycin resistant plastids from the N. plumbaginifolia line Np-LR400-Hyg1. Clones derived from protoplast fusion were selected by kanamycin and hygromycin resistance, transgenic nuclear markers. Somatic hybrids were then grown on a selective streptomycin or lincomycin medium, or in the absence of either drug to a 50 to 100 mg size callus. Southern analysis of a polymorphic region of plastid DNA (ptDNA) revealed that somatic hybrids grown on streptomycin contained almost exclusively ptDNA from the streptomycin resistant parent, somatic hybrids grown on lincomycin contained almost exclusively ptDNA from the lincomycin resistant parent whereas somatic hybrids grown in the absence of either drug contained mixed parental plastids. Sensitive ptDNA was below detection level in most clones on selective medium, but could be recovered upon subsequent culture in the presence of the appropriate drug. The drugs streptomycin and lincomycin provide a powerful selection pressure that should facilitate recovery of plastid transformants.     

In search of mammary gland stem cells

Summary The evidence for mammary epithelial stem cells and their phenotypic characteristics in normal and neoplastic development is reviewed. The presence of stem cells in all parts of the mammary parenchyma at all stages of differentiation has been demonstrated by transplantation experiments. The phenotypic characterization of stem cells has been defined by a battery of monospecific antibodies. These studies suggest that a mammary epithelium stem cell compartment exists in the basal layer of the gland as well as in the end bud. Whether these same stem cells are expressed in mammary preneoplasias and neoplasias has not been answered conclusively. Phenotypic markers specific for stem cells and stably expressed in transformed cell populations are needed to follow the fate of stem cells.Dedicated to Professor Stuart Patton on the occassion of his 70th birthday.     

Evolutionary analysis of Pinus densata (Masters), a putative Tertiary hybrid.

Summary Restriction fragment analysis and heterologous hybridization of chloroplast (cp) DNA was used to develop species-specific markers for P. tabulaeformis, P. yunnanensis and P. massoniana. Fragment patterns created by the BclI and DraI restriction enzymes and hybridization patterns to the psbC and psbD probes were distinctive among the three species. No intraspecific variation was detected with respect to any of the cpDNA markers developed in this study. The cpDNA markers obtained were subsequently used to examine the parentage of P. densata, a putative Tertiary hybrid between P. tabulaeformis and P. yunnanensis. The analysis demonstrated for the first time that P. densata populations accommodate chloroplast genomes of P. tabulaeformis and P. yunnanensis, which strongly supports earlier suggestions of the hybrid origin of this species. It appears that P. densata represents a stabilized natural hybrid that has become adapted to high mountain environments where neither of the parental species can normally grow.     

Origin of the Koreans: a population genetic study.

A population genetic study was undertaken to investigate the origin of Koreans. Thirteen polymorphic and 7 monomorphic blood genetic markers (serum proteins and red cell enzymes) were studied in a group of 437 Koreans. Genetic distance analyses by both cluster and principal components models were performed between Koreans and eight other populations (Koreans in China, Japanese, Han Chinese, Mongolians, Zhuangs, Malays, Javanese, and Soviet Asians) on the basis of 47 alleles controlled by 15 polymorphic loci. A more detailed analysis using 65 alleles at 19 polymorphic loci was performed on six populations. Both analyses demonstrated genetic evidence of the origin of Koreans from the central Asian Mongolians. Further, the Koreans are more closely related to the Japanese and quite distant from the Chinese. The above evidence of the origin of Koreans fits well with the ethnohistoric account of the origin of Koreans and the Korean language. The minority Koreans in China also maintained their genetic identity.     

Molecular tools in marine ecology

Molecular tools have diverse applications in marine ecology. In microbial systems, DNA sequences of rRNA and other genes have identified a variety of novel lineages of bacteria inhabiting marine environments that have resisted traditional culture methods. However, relatively few natural populations have been characterized due to the rather labor-intensive methodologies employed. Recent technological developments such as in situ PCR and flow cytometry promise to greatly enhance the speed at which microbial taxa can be identified and enumerated in field collected water and substrate samples; such advances will allow future work to employ the spatial and temporal field sampling required to monitor the impact of natural and anthropogenic changes in the environment. This approach also holds promise for examining physiological status of field collected cells, garnering information on such elusive parameters as growth rates and the extent of nutrient limitation under natural conditions. Studies of macrobiota have similarly benefited from the use of molecular approaches to species identification. This has been particularly true with regard to distinguishing among larval forms of closely related taxa which are nearly identical morphologically. Genetic variation within species assayed by molecular tools has been useful in examining the stability of populations through time and in assessing patterns of recruitment to geographically separated populations. Enhanced understanding of these ecological problems will also require intensive spatial and temporal monitoring of both larval and adult populations. Often, the newer techniques based on DNA sequence variation have practical advantages over allozyme techniques: e.g., PCR allows assay of minute quantities of DNA that may come from ethanol preserved samples. However, when ample allozyme variation exists to address a given issue, these older techniques may be favored on a variety of criteria, including speed and cost. Hence, choice of methodology should be based on the expected efficiency of a given approach to a specific problem rather than the apparent sophistication of the method itself.     

A MARKER-BASED METHOD FOR INFERENCES ABOUT QUANTITATIVE INHERITANCE IN NATURAL POPULATIONS

A marker-based method for studying quantitative genetic characters in natural populations is presented and evaluated. The method involves regressing quantitative trait similarity on marker-estimated relatedness between individuals. A procedure is first given for estimating the narrow sense heritability and additive genetic correlations among traits, incorporating shared environments. Estimation of the actual variance of relatedness is required for heritability, but not for genetic correlations. The approach is then extended to include isolation by distance of environments, dominance, and shared levels of inbreeding. Investigations of statistical properties show that good estimates do not require great marker polymorphism, but rather require significant variation of actual relatedness; optimal allocation generally favors sampling many individuals at the expense of assaying fewer marker loci; when relatedness declines with physical distance, it is optimal to restrict comparisons to within a certain distance; the power to estimate shared environments and inbreeding effects is reasonable, but estimates of dominance variance may be difficult under certain patterns of relationship; and any linkage of markers to quantitative trait loci does not cause significant problems. This marker-based method makes possible studies with long-lived organisms or with organisms difficult to culture, and opens the possibility that quantitative trait expression in natural environments can be analyzed in an unmanipulative way.     

INFERENCES ABOUT QUANTITATIVE INHERITANCE BASED ON NATURAL POPULATION STRUCTURE IN THE YELLOW MONKEYFLOWER,MIMULUS GUTTATUS

We used a nonmanipulative, marker-based method to study quantitative genetic inheritance in two habitats of a common monkeyflower population. The method involved regressing quantitative trait similarity on marker-estimated relatedness between individuals sampled in the field. We sampled 300 adult plants from each of two transects, one along a stream habitat and another through a meadow habitat. For each plant we measured 10 quantitative characters and assayed 10 polymorphic isozyme loci. In the meadow habitat, relatedness of plants within 1 m was moderate (r = 0.125, corresponding to half-sibs) as was actual variance of relatedness (V_r = 0.044). Significant heritabilities of 50–70% were found for corolla width and the fitness characters of flower number and plant weight. Genetic correlations were strongly positive, but sharing of environmental effects within 1 m was weak. In the stream habitat, levels of relatedness were lower and similar heritabilities were indicated. To detect dominance variance and the correlation of phenotypes due to shared inbreeding, we also estimated higher-order coefficients of relationship and inbreeding, but these did not significantly differ from zero. Laboratory-based estimates of heritability in the field were lower than the marker-based estimates, indicating that natural heritabilities and genetic correlations may be stronger than indicated by controlled studies.     

Identification of genetic factors for alachlor tolerance in maize by molecular markers analysis

Genetic factors controlling tolerance to the herbicide Alachlor in maize were localised by means of two different strategies. In the first approach, backcross (BC) plants, derived from pollen which had been subjected to selective pressure for resistance to the herbicide, were analysed for segregation distortion at 47 RFLP loci and compared to BC plants obtained from non-selected pollen. Preferential transmission of five chromosomal regions where putative QTLs (Quantitative Trait Loci) are localised was revealed in the BC plants from selected pollen. A second approach was based on a classical linkage analysis for segregation of the same set of RFLPs and factors controlling the trait, in a BC population of 210 individuals, by means of regression analysis. This study detected seven significant loci in four genomic regions. Overall, two loci revealed both segregation distortion and association with the expression of the trait, indicating linkage to genes expressed in both gametophytic and sporophytic phase. Three chromosomal regions appeared to carry factors involved in plant tolerance to Alachlor which are not expressed in pollen. Conversely, three loci were linked to factors selectable in pollen, but did not reveal significant association with tolerance in the plant in the segregating populations.     

五种野生稻叶绿体DNA多态性研究

对野生稻 5个种的18个材料的叶绿体DNA(cpDNA)进行了EcoRI的RFLP分析。 结果显示,共有10种酶切模式,不同种野生稻的cpDNA的RFLP类型都不同,而且在其中一些 种内也有变化,尤以O.rufipogon的种内多态性最为显著,并主要与地理来源有关。本研究还在O.punctata的材料中发现一种以往的分析都不曾描述过的多态性模式。通过对结果的分析,探讨了不同种类野生稻的叶绿体基因组之间以及它们与核基因组之间的进化关系。 Abstract:The polymorphisms of chloroplast DNA from 18 materials of 5 wild rice species were investigated using RFLP analysis.10 restriction patterns were obtained from the analysis of these materials.Different species had different of its RFLP patterns chloroplast DNA,and the polymorphisms existed even with species,especially in O.rufipogon varieties of different geographical origins.In O.punctata a new type of rice chloroplast DNA restriction pattern was discovered which had not been reported before.According to the results obtained,the evolutionary relationships among chlorplast genomes,and between chloroplast and nuclear genomes in different wild rice are discussed.     

Introgression of the avian naked neck gene assisted by DNA fingerprints

Theoretical predictions suggest that DNA markers can be useful tools for genomic selection in gene introgression programmes. An experiment was carried out to evaluate the efficiency of using multi-locus DNA markers in an introgression programme designed to transfer the naked neck gene from a donor to a recipient chicken line. The donor line was a commercial egg layer chicken stock heterozygous at the naked neck locus (Na/na⁺), while the recipients were from a Cornish broiler line. These two lines differ markedly in their average body weight, a quantitative trait that can also represent the comprehensive differences between the genomes of the two lines involved. Three groups of naked neck BC₁ individuals were selected according to the following criteria: (i) low band-sharing with their donor grandsires evaluated by multi-locus DNA markers, (ii) high body weight at six weeks of age, and (iii) selection at random as a control group. Birds from each of these groups were mated at random to individuals from the heavier Cornish line to produce three groups of BC₂ individuals whose body weights were recorded weekly from three to seven weeks of age. Results indicated that BC₂ birds obtained from BC₁ parents selected for band-sharing levels and those selected for body weight, performed equally well at 4–7 weeks of age; both were 3.1–3.9% heavier than birds from the randomly selected group. The additional genome recovery of the heavier broiler line, obtained by DNA markers, was found to be in agreement with theoretically predicted values.