embryo doraiancy of pinus jeffreyi murr. seed uptake ... · dase activity, and water-holding...

EMBRYO DORAIANCY OF PINUS JEFFREYI MURR. SEED ASAFFECTED BY TEMPERATURE, WATER UPTAKE,

STRATIFICATION, AND SEED COAT 1'2

EDWARD C. STONESCHOOL OF FORESTRY, UNIVERSITY OF CALIFORNIA, BERKELEY, CALIFORNIA

Almost three hundred years ago the English silvi-culturist, John Evlyn (7), recognized the general prob-lem of forest tree seed dormancy. He did not discussdormancy as such, but he did report that fall or earlywinter sowing of acorns, beech-nuts, haws and hollyberries was a better practice than spring sowing. Hewent on to state that ". . . seeds may be prepared forthe Vernal by being barrell'd or potted up in moist sandor earth stratum during the winter at the expirationwhereof you will find them sprouted and being com-mitted to the earth as apt to take as if they had beensown with the most early ... and will not be much con-cern'd with the increasing heat of the season, as such asbeing crude, and unfermented are newly sown in thebeginning of the Spring. ." Thus, here is perhapsthe first published recommendation for artificial strat-ification of dormant tree seed. It is obvious he didnot know why it was effective-as a matter of fact,we do not know why today, even though we haverecognized some of the associated biochemical changes(6, 8, 11, 12, 14).

Since the turn of the century numerous studies onseed dormancy have been reported; see reviews byMIiller (11), Baldwin (1), Crocker (4), and Crockerand Barton (5). Specific reference to pine seed indi-cates that the dormancy mechanism of that seed isnot entirely clear. For example to overcome dor-mancy, at least five different pregermination treat-ments have been recommended. They are: soakingthe seeds in sulfuric acid (17); soaking the seeds inwater at room temperature (9); soaking the seeds inwvater at low temperature (15, 16); cold storage inwet sand or peat moss (2, 3, 13, 23) and removal ofthe seed coat and papery membrane (19).

The most widely used method of increasing seedgermination is the low-temperature moist storagetreatment generally referred to as stratification. Dur-ing this particular type of pretreatment chemicalchanges apparently take place in the seed, after whichthe seed germinates. For example, Eckerson (6)found during stratification of Crataegus mollis seedsthat the fats decreased while there was a progressiveincrease in sugars, acidity, catalase activity, peroxi-dase activity, and water-holding capacity. Similarresults are reported by Pack (14) for seeds of Juni-perus virginiana, J. communis and J. prostrata. Hefound that fats and proteins decreased while sugars,amino acids, catalase activity, and respiration all in-creased during stratification.

On the other hand Koblet (10) found little changein the character of the food reserve in Pinus strobus

1 Received July 3, 1956.2 Work carried on under the University of California

Agricuiltural Experiment Station Research Project 1577.

seeds during stratification. Only a slight increase inreducing sugars, amino acids, and soluble nitrogen wasnoted. From this he concluded that the low tempera-ture treatment facilitated a faster mobilization of thereserves during germination and ". . . that low tem-peratures could effectively stimulate the plasma orupset its equilibrium thereby giving an impulse togrowth."

The major change Mirov (12) found in stratifiedPinus lambertiana seeds was an increased saturationof the fats. His data also suggested a build-up ofauxin in the endosperm during stratification- achange which Haddock (8) subsequently was unableto detect.

Stratification is required by some but not all spe-cies of pine for satisfactory germination. The "WoodyPlant Seed Manual" (21) lists thirteen native speciesof pine as not requiring stratification. Nevertheless,when seeds of many of these thirteen species arestratified they germinate more quickly and at a moreuniform rate than when they are not stratified (2, 3,13, 23). Therefore a difference in degree rather thana difference in kind is indicated in these two groups.The same metabolic pattern might well prevail inboth groups of pine while the level of substrate, auxin,co-factors and inhibitors might be different.

The dormancy of a large number of temperatezone seeds that responded favorably to stratificationcan also be overcome by breaking or removing theseed coat. This has generally been interpreted as in-dicating non-dormant embryos, and the pines areoften placed in this category (4, 5, 11, 23). However,Stone and Duffield (19) have reported that under cer-tain conditions the embryo of Pinus lambertiana wasat least partially dormant.

An attempt by the author in 1949 to evaluate con-flicting reports by Mirov (12), Haddock (8), andStone (18) regarding auxin change during stratifica-tion was only partially successful (unpublished data).Auxin changes did occur during stratification of Pinuslambertiana seed, but it varied from one seed lot tothe next. It was obvious that "standardized" seedwas needed. Therefore, the following year a largeamount of seed was collected from one tree, dried toa moisture content of ten percent, and then stored at50 C in sealed containers. However, even this uni-formly treated seed could not be considered "stand-ardized" because germination behavior changed con-siderably during the first twelve months of storage.Following this discovery it became important to ob-tain more specific information about germination be-havior prior to the continuation of the auxin studies.Consequently a series of germination behavior studieswas undertaken.

93

www.plantphysiol.orgon May 15, 2020 - Published by Downloaded from Copyright © 1957 American Society of Plant Biologists. All rights reserved.

http://www.plantphysiol.org

PLANT PHYSIOLOGY

During the ensuing five years a number of experi-ments were carried out, none of which were very im-portant in themselves. Now, however, an interestingpicture can be drawn from the overall data. Not onlywas the germination followed for Pinus lambertianaseed which normally requires stratification but alsofor Pinus jeffreyi which normally does not (21).Relevant parts of this study that deal with Pinusjeffreyi are reported here.

MIATERIALS AND METHODSThe seed used in this study was collected in 1952

from several trees located in northeastern California.After air drying for a week or more the seed wascleaned, sealed in five gallon cans, and shipped toBerkeley. There it was stored at 16 C until used in1954 and 1955. The moisture content of the seedvaried from 10 to 13 %, with an average of 12 %.Cutting tests showed that 4 % of the seeds wereempty or otherwise morphologically defective. At thetime of this study freshly harvested seed was notavailable. However, with Pinus jeffreyi this does notassume the importance it does with Pinus lambertiana.Seeds of Pinus jeffreyi store well at 10 C for severalyears withouit any appreciable reduction in its germi-nation (21).

Germination techniques, sample size, and the se-lection of appropriate germination temperatures werebased upon preliminary experiments. The germina-tion temperatures selected were 5, 15 and 250 C, andin each case were held to within + 0.5° C throughoutthe experiment. The samples were replicated tentimes and were of ten seeds each.

As a measure of the effect of the various seedtreatments upon germination at the different tempera-tures, the number of days required to reach 50 %germination was selected. The 50 % germinationlevel was used rather than the 90 % or some otherlevel, because germination was close to its maximumrate at this point and because the total germinationof each seed sample used at least reached the 50 %level. For a statistical expression of the significanceof the difference of germination at the 50 % germina-tion level, the standard error (S.E.) of the differenceof the means was calculated and the t-test applied.

In the water uptake experiments, samples oftwenty-five seeds each, replicated five times were used.Although the S.E. of the mean was large the effectof the treatment, e.g., difference in water uptake afterremoval of the seed coat, was always many timesgreater and a statistical expression of the significanceof the difference was not considered necessary.

STRATIFICATION: Two different methods of strati-fication were used. The first, or standard methodconsisted of mixing the seed with wet vermiculite,pouring the mixture into pint ice cream cartons, seal-ing the cartons and then storing them in a cold roomat 5° C for three months.

The second method consisted of first soaking theseed for 48 hours in 50 C tap water through which airwas continuously bubbled. Then the seed was re-

moved, sealed in large test tubes, without vermiculiteor other stratification media, and stored in a coldroom at 50 C for three months.

GERMINATION OF WHOLE SEED: These studies werecarried out using Petri dishes filled with wet vermicu-lite. Ten seeds were placed on the bottom of eachdish before the vermiculite was added. It was thuspossible to observe germination3 without having toremove the seed from the germinating medium. ThePetri dishes were stacked on shelves in the appropri-ate incubator and inspected daily. From time to timewater was added when the surface of the vermiculiteappeared dry. Any water not absorbed by the ver-miculite was readily observed and drained off aftereach addition of water. Except for the short periodswhen measurements were being made, seeds were keptin the dark.

GERMINATION WITHOUT THE SEED COAT: In gen-eral the procedure was the same as outlined by Stoneand Duffield (19). Seeds were surface sterilized bysoaking for five minutes in bromine water, before be-ing opened in a sterile transfer room. Then after theseed coat and papery membrane, derived from theinner portion of the integument, and the remains ofthe nucellus were removed, the embryo, still sur-rounded by the endosperm, was placed upon solidified1 percent agar in 25 x 200 mm screw top culturetubes. These were then placed in the appropriate in-cubator and germination regularly observed. Again,except for the short periods when measurements werebeing made seeds were kept in the dark.

Some contamination did occasionally occur. How-ever, since each seed was in a separate culture tube,contamination did not spread and in these studies wasnever of any consequence.

WATER UPTAKE MEASUREMENTS: These measure-ments were made with: (a) whole seed, (b) seed fromwhich the seed coat and papery membrane had beenremoved, and (c) isolated embryos and endospermtissue. In some experiments the seed was stratified,in others it was not, and in still others the seed waskilled 4 by heating for one-half hour at 1000 C instoppered test tubes.

One of the methods used to follow water uptakewas to place the seed in Erlenmeyer flasks half filledwith water through which air was bubbled. Thenevery twenty-four houirs samples of the seed weretaken out, the seed coat removed and the moisturecontent of the combinedl endosperm and embryo tis-sues determined.

3 Germination, as used in this paper, refers to theinitial appearance of the radicle. Subsequent elongationof both the radicle and the hypocotyl are not in anyway reflected in the germination data as is the casewhen germination is determined by the number of seed-lings appearing above the ground. The need for thisdistinction will be developed in a subsequent paper.

4 When seed was so treated the embryos, contrary toembryo behavior from untreated seed, did not show anygrowth activity when excised. Furthermore there wasno measurable gas exchange in such seeds even afterthey had imbibed water.

94



STONE-EMBRYO DORMANCY OF PINE

Another method was used in following the wateruptake of isolated embryos and endosperm tissue.The particular tissue was placed on a stack of water-saturated filter papers in the bottom of a Petri dish.Saturation of the paper was maintained by the re-peated addition of sterile w-ater and by keeping thecover on at all times except wlhen weighings were be-ing made. Individual seeds w-ere weighed daily on aRoller-Smith balance in a sterile transfer room.Necessarily seeds were removed from the incubatorsfor short periods of time. However, the maximumtime any seed was out during any 24-hour period was5 minutes. The steps associated with sterile removalof the seed coat as outlined earlier were also followedin the water imbibition sttudies with endosperm andembryos in order to reduce contamination and theattendlant complications.

RESULTSGERMINATION OF STRATIFIED AND UNSTRATIFIED

WHOLE SEED: At 250 C, germination of the unstrati-fied seeds reached the 50 % level in 23 days. On theother hand, when stratified, the 50 % level wasreached in three days (fig 1).

A ten-degree drop in the germination temperaturehad a marked effect upon the unstratified seeds, butlittle effect upon the stratified seeds. Fifty-percentgermination of the unstratified seeds required 115days while the stratified seeds reached the same levelin only 6 days (fig 2).

The same general pattern was continued when thegermination temperature was reduced to 50 C, al-though the order of magnitude was different. At thistemperature unstratified seeds took 175 days for 50 %to germinate while stratified seed took only 50 days(fig 3). Thus, stratification reduced the germinationtime at the three temperatures used.

GERMINATION AFTER REMOVAL OF THE SEED COATAND PAPERY MEMBRANE: Removal of the seed coatsand papery membranes from the unstratified seedsincreased the germination rate at all three tempera-tures. At 25° C the number of days required for 50 %of the seeds to germinate was reduced from 23 to 4; at15° C it was reduced from 115 to 14; and at 50 C itwas reduced from 175 to 95 (figs 1 to 3). Thus hereis a pattern similar to that achieved through stratifi-cation. However, although the patterns are similar,stratification is more effective in increasing the ger-mination rate than is the mere removal of the seedcoats and papery membranes. At 250 C, 50 % of thestratified seeds germinated in 2 days, but it took 4days when unstratified seed were used and only theseed coats and papery membranes removed; at 15° Cit took 6 days and 14 days respectively and at 50 C ittook 50 days and 95 days respectively (figs 1 to 3).

When the seeds were stratified and the seed coatsand papery membranes were also removed the mostrapid germination was attained. Once again the ef-fect of stratification was more evident at 50 C than at15° C or 250 C. Even at 50 C germination was fairlyrapid with 50 % of the germination completed in 10

100

Cw

z

0

z

wa.

Ow

z

w0

zwuwa.

80

60

20

so

60

40

20

TRAT. MINUS S.C. AND MEMS.

STRAT WTH SC AN MEMBC',

NSTRAT. MINUS S.C.

NSTRAT. WITH S.C.

0

10 20 50 100

TIME IN DAYS

STRAT. WITH S.C.

UNSTRAT. MINUS S.Co AND MEMB.

STRAT. ANUS S.C AND MEMS.

UNSTRAT. WI

0 10 20 50 100

TIME IN DAYS

lso 200

ITH S.C.

Iso 200

FIG. 1 (top). Germination at 25° C of stratified andunstratified seeds, with and without the seed coats andpapery membranes. The number of days for 50 % ofthe seeds to germinate varied significantly (0.95 level)between treatments.

FIG. 2 (bottom). Germination at 15° C of stratifiedand unstratified seeds, with and without the seed coatsand papery membranes. The number of days for 50 %of the seeds to germinate varied significantly (0.95 level)between treatments.

days. In contrast stratified whole seeds took 50 days,unstratified seeds without seed coats and papery mem-branes took 95 days, and whole unstratified seeds took175 days (figs 1 to 3).

These results summarized in figure 4, suggest thatstratification and seed coat removal affect germinationthrough their action on two different steps involved inthe germination of the seed. They also suggest thatembryo dormancy in Pinus jeffreyi is a relative con-

D

95



PLANT PHYSIOLOGY

60Z 60 tTRAT. WIT S.C.

u Ae

IAJa20

TIME IN DAYSI-40 UN~~~STRAT. WITH S.C

Z15v

Q t TR . MINUS SiC. AND IWEMB6

CM STRAT, VTH SC \

z

i5|\ X STRAT MINUS S*C. ANMEI

w~~~~~~~~

LId

00 20 00 100 1 0 2180

TIME IN DAYS

FIG. 3 (top) . Germinatio'n at 5° C of stratified andunstratified seeds, with and without the seed coats andpapery membranes. The number of days for 50%l ofthe seeds to germinate varied significantly (0.95 level-)between treatments.

FIG. 4 (bottom). Average number of days for 509%of the seeds to germinate at 5, 15 and 250 C. Stratifiedand unstratified seeds, with and without the seed coatsand papery membranes are shown. Difference in treat-ments was significant (0.95 level)' at all temperatues.

dition. Further speculation on these points, however,will be deferred until the effect of water uptake ongermination is considered.

WATER UPTAKE OF SEED WITHOUT THE SEED COATAND PAPERY MEMBRANE: At 25E C water uptake isclosely associated with germination, with rapid wateruptake being followed by rapid germination (fig 5).

As a consequence, to the casual observer, it might ap-pear that water uptake per se limits germination.However, such a conclusion is not justified inasmuchas the data do not allow one to specify what is causeand what is effect. This cannot be done unless thetwo processes can be studied separately.

Fortunately this can be accomplished by simply-lowering the germination temperature to 50 C. Atthis temperature both water uptake and germination

14or

1201

z aoo

. L

WATER UPTAKE CGERMe SEEDS INCLe)?,.

W,T

/>WATER PTAKE (ER^^ SEES EXCL.

/ DGERMT¢N~~~~~~~~~.

401

20

H-zLLIUa:LId0.

I 2 3 4 aTIME IN DAYS

8 7

WATER UPTAKE CGERM. SEEDS

UPTAKE CGERM. SEEDS EXCL.)

GERMINATION

TIME IN WEEKS

FIG. 5 (top). Germination and water uptake (% drywt) at 25' C of unstratified seeds from which the seedcoats and papery membranes were removed. Seeds weregerminated on wet filter paper in closed Petri dishes.

FIG. 6 (bottom). Germination and water uptake (%dry wt) at 5' C of unstratified seeds from which theseed coats and papery membranes were removed. Seedswere germinated on wet filter paper in closed Petri dishes.

96a

loo




still occur. However, germination does not start forat least three months while water uptake starts im-mediately (fig 6).

At both 5 and 250 C there is a rapid uptake ofwater during the first 48 hours. After this initialperiod the seed continues to take up water but at amuch reduced rate (figs 5 and 6).

At 250 C the seeds have reached an average mois-ture content of 68 % after three days, at which pointgermination begins. In contrast, at 5° C the seedstake 7 days to reach the same average moisture con-tent and germination begins 74 days after that (figs 5and 6). Thus water uptake alone does not appear tocontrol germination.

Each of the several tissues making up the seed hasits own water uptake pattern. Therefore, to obtain acomplete picture of water uptake, these individualtissues must be studied both separately and in closeassociation. When each tissue is separated from theseed and placed on water-saturated filter paper wateruptake of that particular tissue can be followed bysubsequent weighings. When this was done each tis-sue showed a rapid initial uptake of water (fig 7).This was true for both the dead tissues of the seedconsisting of the seed coat and papery membrane aswell as for the live tissue consisting of the embryoand endosperm. Even when the endosperm andembryo tissue were killed by heat there was still arapid uptake of water during the first 24 hours al-though this in turn was followed by a slight loss inwater during the next 24 hours (fig 7).

It would thus appear that the initial water uptakeby the seed is the result of imbibitional and osmoticforces. In the dead tissues of the seed only imbibi-tional forces appear to be involved. In the live tis-sues made up of the embryo and endosperm osmoticas well as imbibitional forces are indicated, first be-cause the living cell normally contains considerableamounts of osmotically active substances and secondbecause of the loss of water from the dead embryotissue during the second 24-hour period (fig 7). Sucha loss in water can only be explained if one assumesthat the semipermeable condition of the cell mem-brane is destroyed when the tissue is killed by heat.Under such conditions the osmotically active sub-stances would slowly diffuse out of the tissue into thesurrounding water and as a consequence there wouldbe a concomitant loss in water from the tissue.

After the rapid initial uptake of water by all thetissues-both dead and alive-the live tissues continueto absorb water at a reduced rate until the seed ger-minates (figs 5 and 6). This slow uptake is probablyassociated with a conversion within the cells of os-motically inactive substances such as starch to os-motically active substances such as glucose (5, 6).When the seed germinates there is again rapid uptakeof water which is obviously necessary if the newlyformed dividing cells of the embryo are to elongate.

At 25° C the water uptake pattern is essentiallythe same as at 50 C except that there is a telescopingof the time scale. Both curves show a sharp initial

- I35 IVE EMBRYO

1- 120 l>120~ ~ DEAD

EMBRYO

05 L

fi4S k SEEDCOAToIt30D t~DEAD ENDOSPERM ---

0 2 3 4 S 6 i 8 9 10TIME IN DAYS

90

80

> 70 I

1-60//

30 24 48729

45 /EE COA

X 3

O 24 4 a 72 96

TIME IN HOURS

FIG. 7 (top). Water uptake (% dry wt) of the indi-vidual tissues making up the seed. The various tissueswere held on wet filter paper in closed Petri dishes.

FIG. 8 (bottom). Water uptake (% dry wt) by intactseeds and seeds from which the seed coats and paperymembranes had been removed. Seeds were in tap waterat 5° C through which a stream of air was bubbled.

rise, followed by a gentle rise, followed finally by an-other sharp rise when the seed germinates (figs 5 and6).

WATER REQUIRED FOR STRATIFICATION: Under nor-mal stratification procedures the seed is free to con-tinue water uptake throughout the period of coldtreatment-three months in this case-which it does.This again complicates the interpretation of the rela-tion between germination and water uptake. Is theslow continuous water uptake and resulting high mois-

97



PLANT PHYSIOLOG;Y

ture content responsible for the germination behaviorof the stratified seed or is this behavior the result ofchemical changes within the seed?

Fortunately again, this was easy to determine. In-stead of providing a continuous source of water dur-ing stratification the seeds were soaked 48 hours in50 C water through which air was continuouslybubbled. Then the seeds were taken out, dried with apaper towel, and sealed in a number of large testtubes. These in turn were placed in incubators at5, 15, and 250 C for three months. At the end of thattime the seeds were removed, planted in wet vermicu-lite, and then placed in a 15° C incubator. Undersimilar conditions, seeds stratified by the usual processgerminated readily (50 % in five days) while unstrati-fied seeds required at least 118 days (fig 4). Of thetreated seeds only those held for 3 months at 50 Chad an appreciable germination in less than 60 days.Sixty percent of these seeds germinated in five daysthus indicating that the seeds could indeed be strati-fied by this process. It is therefore apparent that allthe necessary uptake of moisture occurred during thefirst forty-eight hours when the seeds were soaked(fig 8). After that, during the required remainingninety days, chemical changes essential to rapid ger-mination at 50 C apparently occurred provided theseed was kept at 50 C, but not if it was kept at 15 or250 C. Thus again moisture per se does not appearto be limiting.

PERMEABILITY OF THE SEED COAT AND PAPERYMEMBRANE TO WATER: A comparison of water uptakeby intact seeds and seeds divested of their seed coatsand papery membranes, demonstrated the perme-ability of these two layers to water. When these seedswere soaked in 50 C watei through which air was con-tinuously bubbled, rapid water uptake did occur (fig8). It is true that water uptake was more rapidwhen the seed coats and papery membranes had beenremoved. However, since the seeds still failed to ger-minate when the lag in water uptake was finally over-come, it would not appear that interference withwater uptake per se is the factor limiting germina-tion of Pinus jeffreyi seeds.

DISCUSSION AND CONCLUSIONSFrom the data it would appear that a reasonable

working hypothesis for further study of the seeddormancy mechanism in pine can be derived. Basi-cally a two-step process is indicated, such as S .-- I -) Gwhere S is the substrate, I is one or more intermediatecompounds, and G is growth.

First let us examine this hypothesis in light ofgermination behavior at 50 C (fig 3). At this tem-perature removal of the seed coat did not in itselfpromote rapid germination. This only occurred whenthe seeds had been previously stratified. When theseed coats were removed from the unstratified seeds,germination did not begin for at least ninety days.From then on, however, germination was as rapid asif stratified seed had been used. This would be ex-pected if intermediate compounds I accumulated in

the unstratified seed during the initial ninety dayperiod at 50 C. Thus it would appear that at 50 Cthe seed coat does not interfere with the accumulationof I, but does interfere with its utilization in growth.

Germination of stratified seeds at this low tem-perature was retarded unless the seed coats were re-moved. This suggested interference with either wateruptake or gas exchange or both. Since the seed coatis permeable to water, uptake of water per se wouldnot appear to be limiting. On the other hand, gasexchange is at least slowed down by the presence ofthe seed coat. This would tend to reduce the partialpressure of 02 within the seed which in turn could beexpected to interfere with certain oxidative reactionsnecessary for the utilization of material accumulatedin the seed during stratification such as the postulatedintermediates I.

Thus the proposed reactions S -- I -+ G in whichI slowly accumulates during stratification and in turnis rapidly utilized for growth when the seed coat is re-moved is compatible with the data presented in figure3.

Next let us examine the hypothesis in light of ger-mination at 15° C (fig 2). Here again both seed coatremoval and previous stratification at 50 C increasedthe germination rate. However, there was one princi-pal difference. Germination at 50 C, even though theseed coats had been removed, took place only aftersufficient time had elapsed for the seeds to becomestratified-aproximately ninety days at 50 C for Pinusjeffreyi. At 150 C this was not true. Jeffrey pineseeds cannot be "stratified" at this higher temperature.Furthermore, removal of the seed coats from unstrati-fied seeds, where S -e I would not have been com-pleted, speeded up germination by one-hundred dayswhile removal of the seed coats from stratified seedswhere S -e I would already have been completed ger-mination was only accelerated by five days. Thus itwould appear that at this temperature, unlike at 50 C,the seed coat interferes primarily with the S -- I step.

If at 150 C S -) I goes on rapidly once the seedcoat has been removed, what then can be the explana-tion for the retarding effect of the seed coat? Onceagain the partial pressure of 02 in the seed must beconsidered. It is possible that in the presence of theseed coat and at this higher temperature, with theassociated increase in respiratory activity, the partialpressure of 02 in the seed is reduced to a criticallylow level where S is converted to other compounds Cinstead of being converted to I. If this occurred theremight not be enough I accumulate for subsequentgrowth and germination, e.g., S -- I -+ G. In effect

Cthis would involve reactions of the same general typeas those associated with the Pasteur effect (20). Foradditional speculation on the role of oxygen in thegermination of dormant and stratified seed, the recentbrief review article by Vegis (22) is suggested.

These data indicate the advantages of studyinggermination at different temperatures. The retarding

98




effect of the seed coat can be stuidied at 150 C whereit appears to interfere with the accumulation of inter-mediates. On the other hand at 50 C the effect of theseed coat on the utilization of the intermediates forgrowth can be studied. Also at 50 C the accumula-tion of intermediate I and its chemical identificationcan be most profitably studied.

Thus we have the basis from studies here reportedupon for proposing a working hypothesis and also thebasis for investigating its validity.

SUMMARY1. At 25 and 150 C removal of the seed coats and

the papery membranes from unstratified Pinus jef-freyi seeds allowed rapid germination while at 50 Cit did not. However, when the seed coats and paperymembranes were removed from stratified seeds, ger-mination was rapid at 5 as well as at 15 and 25° C.This was interpreted as indicating a condition of"relative" embryo dormancy which was overcome bystratification.

2. The seed coat and papery membrane sur-rounding the endosperm were found to be permeableto water. Furthermore, "stratification" was accom-plished with only that amount of water absorbed bythe seed during its first 48 hours in contact withwater. Thus water uptake per se did not appear tolimit germination.

3. Water uptake and germination were readilyseparable at 5 but not at 250 C.

4. The data suggested a convenient workinghypothesis for the continued study of the seed dor-imancy mechanism in pine. In effect a two stepprocess was indicated such as S -+ I -+ G where S isthe substrate, I is one or more intermediate com-pounds, and G is growth.

5. The effect of the seed coat on germination at15 and 25° C suggests reactions of the same generaltype as those associated with the Pasteur effect.

LITERATURE CITED1. BALDWIN, H. I. Forest Tree Seed. 240 pp. Chron-

ica Botanica, Waltham, Massachusetts. 1942.2. BARTON, L. V. Hastening the germination of south-

ern pine seeds. Jour. Forestry 26: 775-785. 1928.3. BARTON, L. V. Hastening the germination of some

coniferouis seeds. Plant Physiol. 17: 88-117. 1930.

4. CROCKER, W. Growth of Plants. 459 pp. Reinhold,New York. 1950.

5. CROCKER, W. and BARTON, L. V. Physiology ofSeeds. 267 pp. Chronica Botanica, Waltham,Massachusetts. 1953.

6. ECKERSON, S. A. A physiological and chemical studyof after ripening. Bot. Gaz. 55: 286-299. 1913.

7. EVLYN, J. Silva. A discourse of forest trees andthe propagation of timber. 83 pp. Martyn andAllestry, London. 1664.

8. HADDOCK, P. G. A study of the rest period in seedsand buds of P. Lambertiana Dougl. and P. JeffreyiMurr. Ph.D. thesis, Univ. of California. 1942.

9. JACOBS, A. W. Hastening the germination of sugarpine seeds. Jour. Forestry 23: 919-931. 1925.

10. KOBLET, R. Vber die Keimung von Pinus strobusunter besonderer Birucksichtigung der Herkumftdes Samens. Ber. schweiz. botan. Ges. 41: 199-283. 1932.

11. MILLER, E. C. Plant Physiology, 2nd ed. 1201 pp.McGraw-Hill, New York. 1938.

12. MIRov, N. T. The relations of some internal factorsto germination of seeds of P. jeffreyi Murr. andP. lambertiana Dougl. Ph.D. thesis, Univ. of Cali-fornia. 1936.

13. MIROV, N. T. A note on germination methods forconiferous species. Jour. Forestry 34: 719-723.1936.

14. PACK, D. A. Chemistry of after ripening, germina-tion and seedling development of Juniper seeds.Bot. Gaz. 71: 32-60. 1921.

15. RUDOLF, P. 0. Cold soaking-a short cut substitutefor stratification. Jour. Forestry 48: 31-32. 1950.

16. RUDOLF, P. 0. Cold soaking jack pine. Jour. For-estry 50: 626. 1952.

17. SHOW, S. B. Methods of hastening germination ofsugar pine seed. Jour. Forestry 15: 1003-1006.1917.

18. STONE, E. C. Auxin and respiration changes duringstratification of sugar pine, P. lambertiana Dougl.seed and their relation to subsequent embryogrowth. Ph.D. thesis, Univ. of California. 1948.

19. STONE, E. C. and DUFFIELD, J. W. Hybrids of sugarpine by embryo culture. Jour. Forestry 48: 200-201. 1950.

20. TURNER, J. S. Respiration. Ann. Rev. PlantPhysiol. 2: 145-168. 1951.

21. U. S. FOREST SERVICE. Woody Plant Seed Manual.416 pp. U.S. Dept. Agr. Misc. Publ. 654. 1948.

22. VEGIS, A. Formation of the resting condition inplants. Experientia 12: 94-99. 1955.

23. W7AKELY, P. C. Some observations on southern pineseed. Jouir. Forestry 29: 1150-1164. 1931.

99



embryo doraiancy of pinus jeffreyi murr. seed uptake ... · dase activity, and water-holding...

Documents