NUCLEOLAR VARIATION DURING DIFFERENTIATION OF PHLEUM ROOT EPIDERMIS

Striking differences in nucleolar volume were found between trichoblasts and hairless initials all along the differentiation gradient of the tissue. The larger nucleoli of the trichoblasts were evident from the onset of their differentiation in the meristem and remained so throughout the growing 1000μ of epidermis. At the same time, nucleoli of the alternating, hairless initials rapidly became reduced in volume and virtually disappeared in maturing cells. Nuclear volume was relatively constant throughout the epidermis, so that nuclear:nucleolar volume ratios clearly indicated the nucleolar differences between the shorter trichoblasts and longer hairless initials. Along with the first signs of nucleolar volume difference in the meristem, there were higher concentrations of RNA and ribonucleoprotein in nucleoli and cytoplasm of trichoblasts compared with hairless initials. Although these chemical differences occurred principally in meristem cells, protein content was much higher in trichoblasts than in the alternating hairless initials of the enlargement zone, 300–500μ from the root tip apex. There was no essential difference in protein content between the 2 cell types in the meristem or in older enlarging cells. The data suggested that the initial increase in nucleolar volume and content of the meristem trichoblasts led to their increased protein content and enzyme activities during the enlargement phase of their differentiation. The sharp reduction in nucleoli of the hairless initials, at the same time, led to their generally lower metabolism during tissue differentiation.     

Changes in cell length and mitotic index in vascular pattern-related pericycle cell types along the apical meristem and elongation zone of the onion root

Summary Three pericycle cell types (opposite xylem, opposite phloem and intervening) distinguished by their location in relation to different elements of the vascular system were studied in the adventitious root ofAllium cepa L. Changes in cell length and mitotic index were analysed in these cells along the apical meristem and elongation zone of the root. The opposite phloem and intervening pericycle cells are significantly shorter than the opposite xylem pericycle cells in the apical half of the meristem. Between 1,200 and 1,400 m behind the tip, length became similar in all three pericycle cell types, while in more proximal zones the opposite phloem cells were significantly longer. These results suggest that the number of transverse divisions is different in the three types of pericycle cells. In the apical half of the meristem, mitotic index increased in intervening and opposite xylem cells but remained unchanged in opposite phloem cells, a fact likely to account for the relative lengthening of the latter. In the proximal half of the meristem, mitotic index fell in all three cell types until cell division had ceased. However, mitotic index in opposite xylem cells remained high for longer than in the other two cell types, implying that increase of the mean cell length in the former was slower. These results suggest that differences in mean cell length between the three pericycle cell types are due to different rates of proliferation.     

Anatomy of Jojoba (Simmondsia chinensis) seed and the utilization of liquid wax during germination

Much work has been done on the agricultural potential of Jojoba, but little on the anatomy of the mature plant or seed. Our investigations concern the structure of the embryo of mature seeds and their external morphology during early germination. The embryo is straight and investing. A hypocotyl sheath surrounds the radicle like a hollow cone. The apical meristem is a low mound of cells in a shallow depression between the broad short petioles of the cotyledons. During germination these petioles lengthen and force the embryo away from the coytledons and seed coat. The hypocotyl elongates and the primary root rapidly extends and is well developed before the apical meristem becomes active. A mature imbibed seed contains approximately fifty percent liquid wax. After germination there is a linear decrease in the amount of wax to approximately ten percent at thirty days.     

The mitochondrial cycle of Arabidopsis shoot apical meristem and leaf primordium meristematic cells is defined by a perinuclear tentaculate/cage-like mitochondrion

Plant cells exhibit a high rate of mitochondrial DNA (mtDNA) recombination. This implies that before cytokinesis, the different mitochondrial compartments must fuse to allow for mtDNA intermixing. When and how the conditions for mtDNA intermixing are established are largely unknown. We have investigated the cell cycle-dependent changes in mitochondrial architecture in different Arabidopsis (Arabidopsis thaliana) cell types using confocal microscopy, conventional, and three-dimensional electron microscopy techniques. Whereas mitochondria of cells from most plant organs are always small and dispersed, shoot apical and leaf primordial meristematic cells contain small, discrete mitochondria in the cell periphery and one large, mitochondrial mass in the perinuclear region. Serial thin-section reconstructions of high-pressure-frozen shoot apical meristem cells demonstrate that during G1 through S phase, the large, central mitochondrion has a tentaculate morphology and wraps around one nuclear pole. In G2, both types of mitochondria double their volume, and the large mitochondrion extends around the nucleus to establish a second sheet-like domain at the opposite nuclear pole. During mitosis, approximately 60% of the smaller mitochondria fuse with the large mitochondrion, whose volume increases to 80% of the total mitochondrial volume, and reorganizes into a cage-like structure encompassing first the mitotic spindle and then the entire cytokinetic apparatus. During cytokinesis, the cage-like mitochondrion divides into two independent tentacular mitochondria from which new, small mitochondria arise by fission. These cell cycle-dependent changes in mitochondrial architecture explain how these meristematic cells can achieve a high rate of mtDNA recombination and ensure the even partitioning of mitochondria between daughter cells.     

What the Distribution of Cell Lengths in the Root Meristem Does and Does Not Reveal About Cell Division

Roots have long been realized to be useful material for studies of cell division. Despite this long history of use, the behavior of cells in the meristem is often misinterpreted. A common error is to argue that differences in cell length reflect differences in cell division rate. In this article we explain the fallacy behind this argument and show how the analysis of cell length distribution can lead to insight about the root meristem. These observations support a model for the root meristem where cells of various tissues grow at the same relative growth rate and divide at the same frequency, indicating that these growth parameters are built into the cells at a fundamental level. The differences in cell length between various tissues appear to arise at their formation, first at the tissue initials and ultimately in the seed. Length differences among mature cells may be enhanced by differences in the location within the meristem where division ceases. Discovering mechanisms regulating the length of initial cells and the position where cells cease division requires a realistic understanding of how growth constrains the division behavior of dividing cells.     

锦橙汁囊的超微结构

用常规电镜方法观察了锦橙［Ｃｉｔｒｕｓｓｉｎｅｎｓｉｓ （Ｌ．） Ｏｓｂ．］汁囊从原始细胞到发育为一个具柄的成熟汁囊的过程中,汁囊构成细胞超微结构的变化。锦橙汁囊原始细胞及发育为球状体时的构成细胞以及柱状结构顶端的细胞都是一种典型的分生组织细胞。在细胞质中有包括线粒体、质体、内质网、核糖体等丰富的细胞器,但没有观察到高尔基体。这些分生细胞分裂一段时期后就停止活动,逐渐分化为适应贮藏功能的液泡化薄壁细胞。分生细胞开始分化时,在细胞中出现许多小液泡和高尔基体。这些小液泡逐渐地融合,同时细胞质变少,最后形成一个有中央大液泡的薄壁细胞,在紧贴细胞膜的薄薄的一层细胞质中有线粒体、质体、高尔基体以及含有许多脂滴的杂色体。但成熟果实中汁囊的薄壁细胞中几乎没有任何细胞器。     

Mitotic activity and cell elongation in geostimulated roots of Zea mays

Mitotic activity was investigated in the primary meristem of horizontally oriented excised root tips of Zea mays during the first six hours of their georeaction. The only statistically significant change that could be detected in the meristem was a decrease of the length of its upper half. No significant difference in mitotic activity was found between the upper and lower halves of roots kept continuously horizontal for 6 h. Cell proliferation thus seems relatively insensitive to changes in the redistribution of endogenous growth regulators that are believed to occur within the meristem during the onset of geotropism. In the zone of bending proximal to the meristem cell length was significantly greater in the upper half than in either the lower half or in the equivalent position in vertical control roots. Thus, cell elongation seems to be promoted in the upper half of the horizontal root. Thus, The differences in cell length were not accompanied by any change in the proportion of nuclei synthesising DNA in these elongating, non-meristematic cells.     

Effects of PVA and PEG Pretreatment on Development and Ultrastructure of Plumular Root Mitochondria in Soybean Seed During Low Temperature Imbibition Process

Electron microscope observation revealed that mitochondria in plumular root cells of soy bean seed were invisible at the beginning of seed germination. They were sequentially formed re-differentiated and developed from proplastids during the process of germination. The proplastids in the control plumular root markedly expanded at low temperature imbibition process and did not develop into mitochondria when normal temperature was restored. Even the structure of reticulata membrane was further destroyed and vacuolized. In contrast, mitochondria of soybean seed pretreated with polyvinyl alcohol (PVA) and polythylene glycol-6000 (PEG) continused to develop well under the same conditions, as characterized by a clear structure of bilayered membrane and tube cristae. The results suggest that continuous re-differentiation and development of proplastids into mitochondria at low temperature imbibition process play an important role in the increase of soybean seed vigour and its resistance to chilling injury.     

The seminal root primordia in barley and the participation of their non-meristematic cells in root construction

In addition to the primary seminal primordium, the so-called secondary seminal root primordia are also initiated in a barley embryo. The primary root primordium is developmentally most advanced. It is formed by root meristem covered with the root cap, and by a histologically determined region with completed cell division. On germination, the restoration of growth processes begins in this non-meristematic region of root primordium by cell elongation, with the exception of the zone adjacent to the scutellar node, the cells of which do not elongate but continue differentiating. In the root primordia initiated later, the zone with completed cell division is relatively shorter, in the youngest primordia the non-meristematic cells may be lacking. The root meristem is reactivated after the primary root primordium has broken through the sheath-like coleorrhiza and emerges from the caryopsis as the primary root. The character of root meristem indicates a reduced water content at the embryonic development of root primordium. With progressing growth the root apex becomes thinner, the meristematic region becomes longer, and the differences in the extent of cell division between individual cell types increase. — The primary root base is formed of cells pre-existing in the seminal root primordium. Upon desiccation of caryopsis in maturation, and subsequent quiescent period, their development was temporarily broken, proceeding with the onset of germination. The length of this postembryonically non-dividing basal zone is different in individual cell types. The column of central metaxylem characteristic of the smallest number of cell cycles, has, under the given conditions, a mean length of about 22 mm, whereas the pericycle, as the tissue with most prolonged cell division, has a mean length of about 6 mm. In the seminal root primordia initiated later the non-dividing areas are relatively shorter. The basal region of seminal roots thus differs in its ontogenesis from the increase which is formed “de novo” by the action of root meristem upon seed germination.     

Morphological differentiation of mitochondria in the early chick embryo: a stereological analysis

The morphological evolution of mitochondria in three cell types of chick embryo in neurulation was analyzed by stereological methods. Mitochondria, showing a random distribution, were characterized by moderate electron-dense matrices and normal cristae. The numerical density of mitochondria significantly increased in the neuroectoderm and epiblastic cells while their volume density remained unchanged. The mitochondria in mesoderm cells were ellipsoidal (axial ratio 2:1) at stages 5 and 8 although they underwent an elongation in neuroectoderm and epiblastic cells (axial ratio from 2:1 to 1.6:1). The individual size of "average mitochondria" in the mesoderm cells was smaller than in other cell types. The total V/S (volume/surface) ratio of mitochondria decreased during neurulation. These morphological changes have been discussed emphasizing the possible metabolical role of mitochondria during morphogenesis.     

The Fine Structure of the Quiescent Centre and Neighbouring Tissues3

The quiescent centre of the root meristem of Zea provides asource of undiffer-entiated, non-meristematic cells whose finestructure we have compared with that of the meristematic, non-differentiatedcells of the meristem and that of the differentiated, non-meristematiccells of the cap. We have shown an association between the developmentof the endoplasmic reticulum and the development of cell walls.Differentiation in the cap is accompanied by a decrease in thenumber of mitochondria and plastics per unit volume of cytoplasmand an increase in their number per cell. There is an even greaterincrease in the number of Golgi bodies per cell. The structureof the mitochondria becomes more elaborate in differentiatingcells especially in the cap and the Golgi increase their sizein meristematic and differentiating cells. These differencesare discussed in relation to some of the views about radiosensitivityand organization.     

Correlations Between the Dimensions of Different Zones of Grass Root Apices, and their Implications for Morphogenesis and Differentiation in Roots

The sizes of different zones within root apices of nine speciesof grass were estimated, and statistically significant correlationswere found between certain of them. The volume of the cap isrelated to the volume of the meristem of the root proper. Thecortical and stelar portions of the meristem are also related,and their lengths and volumes correlate with the volume of thequiescent centre. The volume of the quiescent centre also correlateswith the length of the zone in which periclinal divisions arefound in the inner cortex; these divisions generate the rowsof cells across the cortex. Diameter of the procambial cylinder,quiescent centre volume and vascular complexity are related,though from correlations alone it is not possible to say whetherone of these characters directly influences another as has beensuggested by other workers. All the zones within the root apexprobably form a tightly-integrated developmental unit. Root structure seems to be independent of cell size, thoughcell size correlates with nuclear DNA content which is a species-specificfeature. Gramineae, meristems, morphogenesis, root apices     

Mitochondrial growth and division during the cell cycle in HeLa cells

The growth and division of mitochondria during the cell cycle was investigated by a morphometric analysis of electron micrographs of synchronized HeLa cells. The ratio of total outer membrane contour length to cytoplasmic area did not vary significantly during the cell cycle, implying a continuous growth of the mitochondrial outer membrane. The mean fraction of cytoplasmic area occupied by mitochondrial profiles was likewise found to remain constant, indicating that the increase in total mitochondrial volume per cell occurs continuously during interphase, in such a way that the mitochondrial complement occupies a constant fraction( approximately 10-11(percent)) of the volume of the cytoplasm. The mean area, outer membrane contour length, and axis ratio of the mitochondrial profiles also did not vary appreciably during the cell cycle; furthermore, the close similarity of the frequency distributions of these parameters for the six experimental time-points suggested a stable mitochondrial shape distribution. The constancy of both the mean mitochondrial profile area and the number of mitochondrial profiles per unit of cytoplasmic area was interpreted to indicate the continuous division of mitochondria at the level of the cell population. Furthermore, no evidence was found for the occurrence of synchronous mitochondrial growth and division within individual cells. Thus, it appears that, in HeLa cells, there is no fixed temporal relationship between the growth and division of mitochondria and the events of the cell cycle. A number of statistical methods were developed for the purpose of making numerical estimates of certain three-dimensional cellular and mitochondrial parameters. Mean cellular and cytoplasmic volumes were calculated for the six time-points; both exhibited a nonlinear, approx. twofold increase. A comparison of the axis ratio distributions of the mitochondrial profiles with theoretical distributions expected from random sectioning of bodies of various three-dimensional shapes allowed the derivation of an "average" mitochondrial shape. This, in turn, permitted calculations to be made which expressed the two-dimensional results in three-dimensional terms. Thus, the estimated values for the number of mitochondria per unit of cytoplasmic volume and for the mean mitochondrial volume were found to remain constant during the cell cycle, while the estimated number of mitochondria per cell increase approx. twofold in an essentially continuous manner.     

菜蛾盘绒茧蜂主要寄生因子导致的寄主小菜蛾幼虫脂肪体结构的变化

在不同的寄生状态下,菜蛾盘绒茧蜂Cotesia plutellae不同的寄生因子可引起寄主小菜蛾Plutella xylostella幼虫脂肪体结构发生相应的改变。显微和亚显微形态结构显示： 假寄生后多分DNA病毒和毒液对脂肪体结构的完整性没有显著影响,但细胞内脂质体变得小而密集,线粒体和内质网丰富,并有糖原积累; 正常寄生后,脂肪体结构被破坏,多数线粒体内嵴紊乱,脂质体也变得不规则,特别是当幼蜂完成在寄主体内发育时,寄主体内几乎无完整脂肪体存在。与此同时,同批未被寄生的小菜蛾幼虫发育到4龄末期时,体内脂肪体细胞发育正常,已开始向蛹期细胞形态转化,细胞内脂质体很大,细胞器数量较多、糖原积累丰富, 而且部分细胞已成为游离态细胞。由此证明,寄生蜂携带的寄生因子,如多分DNA病毒、毒液、畸形细胞和幼蜂等,均对寄主脂肪体结构的改变产生影响,但程度明显不同。     

Mitochondrial biogenesis during germination in maize embryos

Mitochondrial biogenesis and metabolism were investigated during maize (Zea mays) seed germination. Mitochondria from dry and imbibed seed exhibited NADH-dependent O(2) uptake that was completely inhibited by KCN and antimycin A. Mitochondria in the dry seed had a lower rate of succinate-dependent O(2) uptake relative to that measured in imbibed and germinated seed. The activities of the tricarboxylic acid (TCA) cycle enzymes, pyruvate dehydrogenase complex, 2-oxoglutarate dehydrogenase complex, NAD-malic enzyme, and citrate synthase, are similarly low in mitochondria from dry seed and this correlates with a lower relative abundance of the mitochondrial matrix-located citrate synthase and pyruvate dehydrogenase complex E1alpha-subunit polypeptides. Electron microscopy revealed that mitochondria in the dry seed have a poorly developed internal membrane structure with few cristae; following 24 h of germination the mitochondria developed a more normal structure with more developed cristae. The mitochondria from maize embryos could be fractionated into two subpopulations by Suc density gradient centrifugation: one subpopulation of buoyant density equivalent to 22% to 28% (w/w) Suc; the other equivalent to 37% to 42% (w/w) Suc. These two subpopulations had different activities of specific mitochondrial enzymes and contained different amounts of specific mitochondrial proteins as revealed by western-blot analysis. Both subpopulations from the dry embryo were comprised of poorly developed mitochondria. However, during imbibition mitochondria in the heavy fraction (37%-42% [w/w] Suc) progressively acquired characteristics of fully functional mitochondria found in the germinated seedling in terms of structure, enzymic activity, and protein complement. In contrast, mitochondria in the light fraction (22% to 28% [w/w] Suc) show no significant structural change during imbibition and the amounts of specific mitochondrial proteins decreased significantly during germination.     

Variation of mitochondrial size during the cell cycle: A multiparameter flow cytometric and microscopic study.

BACKGROUND: Changes in mitochondrial structure and size are observed in response to alterations in cell physiology. Flow cytometry provides a useful tool to study these changes in intact cells. We have used flow cytometry and digital fluorescence microscopy to analyze the variations in mitochondrial size in relation to specific phases of the cell cycle. METHODS: Supravital staining of rat fibroblasts was done with Hoechst 33342 and rhodamine 123, and cells were analyzed in a dual-laser flow cytometer. Synchronized cells at various stages of the cell cycle were analyzed for changes in mitochondrial size. These cells were also examined by electron microscopy, digital fluorescence microscopy and computerized image analysis to compare the lengths of the mitochondria. RESULTS: By using fluorescence pulse width analysis, we observed two populations of mitochondria in intact cells. The percentage of cells with small and large mitochondria at specific stages of the cell cycle indicated that mitochondrial size increases during the cell cycle; early G1 phase cells had the smallest mitochondria and the mitotic phase cells had the largest mitochondria. These results were confirmed by microscopic analysis of cells. CONCLUSIONS: Flow cytometry can distinguish the relative mitochondrial size in intact cells, and in combination with digital microscopy it can be used to study mitochondrial variation during the cell cycle.     

FINE STRUCTURE STUDIES OF PHLEUM ROOT MERISTEM CELLS. I. MITOCHONDRIA

Avers , Charlotte J. (Douglass Coll., Rutgers—The State U., New Brunswick, N.J.) Fine structure studies of Phleum root meristem cells. I. Mitochondria. Amer. Jour. Bot. 49(9): 996–1003. Illus. 1962.—Mitochondrial numbers were computed from thick sections embedded in paraffin and stained with iron hematoxylin, from methacrylate embedded, 0.5 μ-thick sections stained with osmium, and from electron micrographs of ultrathin sections fixed in KMnO₄. The mean mitochondrial counts per average cell (20 × 20 × 10μ) were: 121 ± 4 in paraffin sections, 600 ± 40 in methacrylate sections, and 991 ± 96 from electron micrographs. These numbers were higher than the Janus green B count of 91 ± 2 per cell found during an earlier study. In each case, arithmetical conversion factors were used to calculate total numbers of mitochondria per cell since section thicknesses were less than that of whole cell length or depth. Determinations of numbers of mitochondria probably varied depending on section thickness and specific staining procedures used, but the higher count from electron micrographs was assumed to be reasonable on a volumetric basis. The cytoplasmic volume of the average cell is about 2500 μ³ and the volume of a single average mitochondrion is about 0.1μ^3. On this basis, the variety of structures and their relatively sparse distribution seems possible despite the apparently high numbers found. In addition to mitochondria, the numbers of various cellular components per unit cytoplasmic area were determined. These data showed that mitochondria were about 6 times more numerous than proplastids, about 4.5 times more numerous than Golgi bodies, and even more frequent when compared with vesicles, dense bodies, or microbodies. No correlation was found between cytoplasmic area and numbers of organelles per unit area. Photographic evidence was presented for the occurrence in plant cells of the hepatic cell, polymorphic “lysosome” described by Ashford and Porter. The controversial nature of the lysosome is discussed.     

The Response of the Primary Root Meristem of Zea mays L. to Various Periods of Cold

Primary roots of maize (Zea mays L.) grown in nutrient solutionat 5?C elongate at about 1% of the rate found at 20?C. The apicalmeristem becomes shorter and shows little proliferative activityat 5?C, but following transfer to 20?C mitoses increase in frequencyand the meristem regrows to its original length. Both the amountby which the meristem shortens and the time for its completeregrowth are related to the period spent at 5?C. The shorteningof the meristem suggests that at the lower temperature the balancewhich normally exists between cell production and differentiationis altered, the latter continuing at a relatively faster ratethan the former. A new, steady-state balance between the twoprocesses is re-established during the recovery period. Themeristem recovers as a result not only of its own mitotic activitybut also through stimulation of cell division in the quiescentcentre. The degree to which the quiescent centre is activated,as judged by its mitotic index and the number of nuclei labelledby feeding with tritiated thymidine, increases as the durationof the preceding cold treatment increases. The close relationshipbetween proliferative activity in the quiescent centre and theminimum length of the meristem following the cold treatmentsuggests that there is communication between these two zoneswhich co-ordinates their respective rates of cell productionand helps to maintain a normal meristem structure. The resultsemphasize the importance of the quiescent centre as a reservoirof cells that can re-establish a meristem rendered non-functionalthrough the impact of unfavourable environmental conditions. Key words: Meristem, quiescent centre, root, temperature, Zea mays     

Chronological transition of mitochondrial morphology in Chlamydomonas reinhardtii (Chlorophyceae) poststationary phase growth1

In Chlamydomonas reinhardtii P. A. Dangeard, mitochondrial morphology has been observed during asexual cell division cycle, gamete and zygote formation, zygote maturation, and meiotic stages. However, the chronological transition of mitochondrial morphology after the stationary phase of vegetative growth, defined as the poststationary phase, remains unknown. Here, we examined the mitochondrial morphology in cells cultured for 4 months on agar plates to study mitochondrial dynamics in the poststationary phase. Fluorescence microscopy showed that the intricate thread‐like structure of mitochondria gradually changed into a granular structure via fragmentation after the stationary phase in cultures of about 1 week of age. The number of mitochondrial nucleoids decreased from about 30 per cell at 1 week to about five per cell after 4 months of culture. The mitochondrial oxygen consumption decreased exponentially, but the mitochondria retained their membrane potential. The total quantity of mitochondrial DNA (mtDNA) of cells at 4 months decreased to 20% of that at 1 week. However, the mitochondrial genomic DNA length was unchanged, as intermediate lengths were not detected. In cells in which the total mtDNA amount was reduced artificially to 16% after treatment with 5‐fluoro‐2‐deoxyuridine (FdUrd) for 1 week, the mitochondria remained as thread‐like structures. The oxygen consumption rate of these cells corresponded to that of untreated cells at 1 week of culture. This suggests that a decrease in mtDNA does not directly induce the fragmentation of mitochondria. The results suggest that during the late poststationary phase, mitochondria converge to a minimum unit of a granular structure with a mitochondrial nucleoid.     

Changes in cell-cycle duration and growth fraction in the shoot meristem of Sinapis during floral transition

The cell-cycle duration and the growth fraction were estimated in the shoot meristem of Sinapis alba L. during the transition from the vegetative to the floral condition. Compared with the vegetative meristem, the cell-cycle length was reduced from 86 to 32 h and the growth fraction, i.e. the proportion of rapidly cycling cells, was increased from 30–40% to 50–60%. These changes were detectable as early as 30 h after the start of the single inductive long day. The faster cell cycle in the evoked meristem was achieved by a shortening of the G1 (pre-DNA synthesis), S (DNA synthesis) and G2 (post-DNA synthesis) phases of the cycle. In both vegetative and evoked meristems, both-the central and peripheral zones were mosaics of rapidly cycling and non-cycling cells, but the growth fraction was always higher in the peripheral zone.Abbreviations G1 pre-DNA synthesis phase - G2 post-DNA synthesis phase - GF growth fraction - M mitosis phase - PLM percentage-labelled-mitoses method - S DNA synthesis phase - TdR thymidine