R. L. Phillips

Plants regenerated from relatively undifferentiated callus cultures possess a vast array of genetic changes. Such variations can result in useful agricultural and horticultural products. For other purposes, however, variations in traits other than those of interest may be undesirable--for example, using cultured cells for genetic engineering. Any steps made toward understanding the basis of tissue culture-induced genetic variation should be helpful in developing a more stable and manipulatable somatic cell system. This review provides a glimpse at the specific kinds of genetic changes encountered among regenerated plants and their progeny. Included among these variations are cytosine methylation alterations of the genome. The repeat-induced point mutation (RIP) phenomenon, reported for filamentous fungi, is invoked to provide a framework to consider the origin of variation in plant tissue cultures.

Abstract Effective utilization of cell and tissue culture methods in Zea mays research requires cultures capable of plant regeneration. These differentiated plants would provide a direct link with conventional genetic and breeding procedures. Maize callus from embryo scutellar tissues was initiated and maintained on MS medium inorganic components, Straus medium vitamins and amino acids, 20 g sucrose and 8 g agar per liter, and 2 mg 2,4‐dichlorophenoxyacetic acid (2,4‐D)/liter. Callus has been maintained subculture every 21 to 28 days and has remained capable of differentiation for 9 months. Regeneration of complete plants was accomplished by subculture of callus to 0.25 mg 2,4‐D/liter for 30 days followed by transfer to 2,4‐D‐free culture medium. At 0.25 mg 2,4‐D/liter numerous curled and wrinkled leaves developed. Approximately 200 complete plants have been differentiated. after transfer to the 2,4‐D‐free medium. Root tip cells from five plants indicated that each had 20 chromosomes. After transplantation to soil, 10 to 15% of the plants survived and grew normally. The optimum embryo age for scutellar callus initiation was 18 days post‐pollination. Hormone combinations such as 1 mg 2,4‐D, 4 mg a‐naphthaleneacetic acid (NAA), and 0.05 mg 6‐(γ,γ‐dimethyl allylamino)‐purine (2iP)/liter may increase the efficiency of scutellar callus initiation.

A heuristic model of irradiance fluctuations for a propagating optical wave in a weakly inhomogeneous medium is developed under the assumption that small-scale irradiance fluctuations are modulated by large-scale irradiance fluctuations of the wave. The upper bound for small turbulent cells is defined by the smallest cell size between the Fresnel zone and the transverse spatial coherence radius of the optical wave. A lower bound for large turbulent cells is defined by the largest cell size between the Fresnel zone and the scattering disk. In moderate-to-strong irradiance fluctuations, cell sizes between those defined by the spatial coherence radius and the scattering disk are eliminated through spatial-frequency filtering as a consequence of the propagation process. The resulting scintillation index from this theory has the form σI2=σx2+σy2+σx2σy2, where σx2 denotes large-scale scintillation and σy2 denotes small-scale scintillation. By means of a modification of the Rytov method that incorporates an amplitude spatial-frequency filter function under strong-fluctuation conditions, tractable expressions are developed for the scintillation index of a plane wave and a spherical wave that are valid under moderate-to-strong irradiance fluctuations. In many cases the models also compare well with conventional results in weak-fluctuation regimes. Inner-scale effects are taken into account by use of a modified atmospheric spectrum that exhibits a bump at large spatial frequencies. Quantitative values predicted by these models agree well with experimental and simulation data previously published. In addition to the scintillation index, expressions are also developed for the irradiance covariance function of a plane wave and a spherical wave, both of which have the form BI(ρ)=Bx(ρ)+By(ρ)+Bx(ρ)By(ρ), where Bx(ρ) is the covariance function associated with large-scale fluctuations and By(ρ) is the covariance function associated with small-scale fluctuations. In strong turbulence the derived covariance shows the characteristic two-scale behavior, in which the correlation length is determined by the spatial coherence radius of the field and the width of the long residual correlation tail is determined by the scattering disk.

A set of oat-maize chromosome addition lines with individual maize (Zea mays L.) chromosomes present in plants with a complete oat (Avena sativa L.) chromosome complement provides a unique opportunity to analyze the organization of centromeric regions of each maize chromosome. A DNA sequence, MCS1a, described previously as a maize centromere-associated sequence, was used as a probe to isolate cosmid clones from a genomic library made of DNA purified from a maize chromosome 9 addition line. Analysis of six cosmid clones containing centromeric DNA segments revealed a complex organization. The MCS1a sequence was found to comprise a portion of the long terminal repeats of a retrotransposon-like repeated element, termed CentA. Two of the six cosmid clones contained regions composed of a newly identified family of tandem repeats, termed CentC. Copies of CentA and tandem arrays of CentC are interspersed with other repetitive elements, including the previously identified maize retroelements Huck and Prem2. Fluorescence in situ hybridization revealed that CentC and CentA elements are limited to the centromeric region of each maize chromosome. The retroelements Huck and Prem2 are dispersed along all maize chromosomes, although Huck elements are present in an increased concentration around centromeric regions. Significant variation in the size of the blocks of CentC and in the copy number of CentA elements, as well as restriction fragment length variations were detected within the centromeric region of each maize chromosome studied. The different proportions and arrangements of these elements and likely others provide each centromeric region with a unique overall structure.

Breeding programs in major crops normally restrict the use of parents to those improved for a variety of traits. Gain from utilizing these good × good crosses appears to be high, and improvements are sufficient to encourage continued breeding within narrow gene pools even though each cycle is expected to lead to reduced genetic variability. These finely tuned programs have gradually limited the amount of new diversity introduced into the breeding gene pool. This breeding strategy has led to a genetic gap where there is a large difference in the favorable gene frequency between the improved and unimproved lines and to a narrowing of genetic diversity within elite gene pools. At the same time, evidence has accumulated in plant breeding programs and long‐term selection experiments in several organisms that the genome is more plastic and amenable to selection than previously assumed. In the barley ( Hordeum vulgare L.) case study reported here, incremental genetic gains were made for several traits in what appears, based on pedigree analysis, to be a narrow gene pool. Given this situation, we call for an examination of the generally held belief that the variation on which selection is based in elite gene pools is provided almost exclusively from the original parents. Classical and molecular genetic analyses have shown that many mechanisms exist to generate variation de novo , such as gene amplification and transposable elements. Accordingly, we put forward the hypothesis that newly generated variation makes an important contribution. We also hypothesize that gene interaction, epistasis, is more important than commonly viewed and that it arises from de novo generated diversity as well as the original diversity.