Anatomia de Una Revolucion

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Copyright 1998 by the Genetics Society of America

Perspectives

Anectdotal, Historical and Critical Commentaries on Genetics

Edited by James F. Crow and William F. Dove

Anatomy of a Revolution

Gerald R. Fink

Whitehead Institute, Massachusetts Institu te of Technology, Cambridge, Massachusetts 02142

WITH this volume, Genet ics annou nces that Ara- 1986). Yet, today there is virtually no m ajor academicinstitution or agrotech company that does not have anbidopsish as joined the SecurityCouncil ofModelArabidopsis group; indeed, many formerly disdainfulGenetic Organisms. These favored few form the stan-

botanists are n ow amon g Arabidop sis most avid prop o-dard to which all other organisms are compared. Like

nents. So, what is responsible for this abrupt turn-the SecurityCoun cil of the Un ited Nations, wher e ther e

around? I believe that the rapid rise of Arabidopsis asis broad geographical representation, the Securitya premier genetic system is a consequence of the fortu-Coun cil of Genetic Or ganisms seeks broad ph ylogenetic

itous conflu ence of th ree factorsscientific, social, an drepresentation. Therefore, the current membership

economic.also includes a virus (lambda), a gram positive bacte-The crucial scientific factors were the concurrent de-rium (Bacillus subtilis), a gram negative bacterium (Esche-

velopmen t of Arabidopsis geno mics1 and the technicalrichia coli), a fungus (Saccharomyces cerevisiae) , a u nicellulartoolbox of modern molecular genetics. Together, thesealga (Chlamydomonas reinhardtii), a worm ( Caenorhabditishave made it possible to make mutants, to clone theelegans), an arthropod ( Drosophila melanogaster), a verte-corresponding gene, and to decipher the function ofbrate (Mus musculus), and humans. The idea is that anthat gene. Genomic advances include the developmentintense concentration on the genetics of one of theof h igh-resolution physical an d genetic maps and therepresentatives provides a window on the biology ofattendan t YAC, BAC, an d lambda libraries. This geno -all the other species in that phylum. Arabidopsis hasmic infrastructure has accelerated all genetic work andemerged as the flowering plant delegate, presaging dra-

forms the backbone o f the global Arabidopsis Sequen c-matic changes in agriculture and the topsy-turvy worlding Project. The goal is to have the complete sequenceof nutritional sciences.of the roughly 100 Mb in the Arabidopsis genome byTo be elected to the Security Council, a delegatethe turn of the century.organism must meet ob jective criteria. It must be amena-

In addition to these advances in genom ics, we alsoble to all the classical genetic movestests of dominance,have a panoply of new genetic techniques. There iscomplementation, and recombination. Itsgenome shou lda con venien t tran sposon tagging system ( H e h l 1994;be sequenced or in the process of being sequenced,Azpir oz -Leeh an an d Fel dman n 1997) that permitsn otand it must be readily transformed with DNA. With allonly the rapid isolation of mutants, but also the easyof th ese techniques in place, geneticists can performacquisition of the corresponding genes. There are alsotheir sacred task: conn ecting genes with their function.useful enh ancer trap and gene trap constructs that facili-Having said all this, elevation to membership on thetate cell and tissue localization for developmental andSecurity Council (as with the permanen t members ofregulatory analysis ( Sun dar esan et al. 1995). But it isthe U .N.) has a stron g historical compo nen t. Buttressedthe development of a rapid transformation protocolby90 yearsof study, Drosophila isun likelyto be rep lacedthat has propelled Arabidopsis to center stage.by another Arthropod. And the inclusion of hum ans

Remarkably, Arabidopsis can be transformed simplysmacks of self-interest on the part of the selection com-by dipping whole plants into a solution of Agrobacte-mittee.rium containing a T-DNA plasmid ( Bech t ol d et al.While Arabidopsisfulfillsth e criteria for mem bership,1993; Ch a n g et al. 1994). Plants are inverted and im-its meteoric rise to council rank is still surprising. As

few as 10 years ago, most botanists shunned this tiny

weed despite th e fact that its small size, short gen eration1ArabidopsisWorld Wide Web electronic resources:h ttp:// geno me.

time, and small genome with little repeated DNA were www.stanford .ed u/ Arabidop sis, htt p:// n asc.life.nott.ac.u k/ cen tre.h tml,[email protected] advertised ( Redei 1975; Pr uit t an d Meyer owit z

Genetics 149: 473477 ( June, 1998)


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474 G. R. Fink

mersed in Agrobacterium, and the combo is placed chromosomal location. Currently, each construct ends

up at a different site, making the results subject to theunder vacuum for a brief period. The Agrobacterium

infiltrates the tissues and d elivers the T-DNA plasmid interpretation that some of the phen otypes may be due

to chrom osomal position effects. Th e recent rep ort ofcontaining the DNA of interest into cells, where it be-

comes stably integrated into the genome. As some of a homologous integration event ( Kempin et al. 1997)

holds out the hope that this impasse may soon be sur-the transformed cells develop into gametes, the seeds

of these plants give rise to stable transformants in the mounted.

In addition to these scientific developments, therenext gen eration. Prior to this discovery, tr ansform ation

in Arabidopsis was a nightmare , requ iring a skilled tech- have also been extrascientific factors, both social andeconomic, that were critical to the transcendence ofnician to establish cell cultures from plants, transform

the cells in culture, and then regenerate them into Arabidopsis. A key social factor was the excellent men-

toring of students by the scientists who initially popu-plants. This painstaking p rocess was slow( 46 m onth s),

cumbersome, and inefficient. More often than not, the lated the field (among them, Fr ed Ausu bel , Gl or ia

Cor uzzi, Ron Davis, El l iot Meyer owit z, and Chr isfew transformants obtained had genetic abnormalities

(sterility, somaclonal variation) that made them useless Somer vi l l e). This founding group of molecular biolo-

gists transmitted to their students the community spiritfor further analysis.

Th e in planta transformation protocol is easy and that emerged from the lambda, E. coli, yeast, worm , an d

Drosophila traditions. Equallyimp ortant, these mentorsavoids all the problems of the cell culture approach.

Without much supervision, even a high school student not only allowed but encouraged their students to take

their successful projectswith th em when they embar kedcan generate large n umbers of fertile transformants.

This procedure is now used routinely to identify genes upon their own indepen dent careers, creating a web of

interacting laboratories. This amicable spirit fosteredby functional complementation. On ce the relevant m u-tation is localized on the physical map (1 cM 200 the rapid dissemination of strains, DNA libraries, tech-

niques, and pr ojects with the consequent rapid accelera-kb), DNA segments contained within that region can

be readily transformed into the plant to identify the tion of research.

Anoth er socialfactor was Cold Spring Harb ors adop -one that suppresses the mutant defect. Th e current

protocol makes it practical to scan across a segment of tion of Arabidopsis as the organism for the plant molec-

ular biology course. The plants rapid growth an d shor t50 kb (1015 genes) and to obtain u nambiguous gene

id en tification in 23 m on th s. gen eration tim e m ake it a n atural for a brief sum mer

course program. The Cold Spring Harbor imprimaturThe Arabidopsis tran sformation system still lacks a

reliable method for obtaining homologous integration. served simultaneously to advertise the organism and to

recruit young scientiststo thisnew and burgeoning field.This system would permit the simple construction of

knockouts and the integration of virtually any DNA seg- With the publication of the book, Arabidopsis ( Meyer o-

wit z an d Somer vil l e 1994), Cold Spring Harbor alsoment at a predetermined chromosomal location without

rearranging the rest of the genome. This shortcoming promoted the codification and dissemination of usefullore and techniques.has not precluded the election of Arabidopsis to the

Security Council, especially since other members, nota- Notwithstanding these scientific and social factors,

thisabrup t turnabout could not h ave happened withoutblyDrosophila and the worm, currentlysuffer from this

limitation. However, there are many fascinating plant timely financial backing. The church regrettably no

longer supports plant science as it did in Mendel stime.phen omena that will be u nderstood onlywhen a homol-

o go u s in t egr at io n syst em is d e ve lo p ed . Fo rt un a te ly, t h e N at io n al Scie n ce Fo u n da tio n ( N SF)

stepped up to promote Arabidopsis. There are manyA good example is gene silencing, a phenomenon in

which the presence of extra copies of a gene introduced program d irectors at NSF who deserve thanks, but it

wasDr. DeLil l Nassir who first sen sed th e zeitgeist andby transformation inactivates both the transgene and

the resident homologous gene ( Fl avel l 1994; Mar - champ ioned Arabidopsis when it was not fashion able to

do so. She used discretionary funds, her wise counsel,t ienssen 1996). Silencing is not simp ly a consequence

of transformation because genes that have duplicated and influence within the federal funding agencies to

get the ball rollingsupporting meetings, identifyingspontaneously can also be silenced ( Bender an d Fink

1995). This homology-depen dent gene inactivation may the movers and the shakers, and making sure that the

best Arabidopsis p roposals were supported. We mayhold the secret as to how the genome defends itself

against rampant gene duplication and the invasion of praise the organism and p rize our geniuses, but the

role of federal supp ort in the emergen ce of Arabidopsisalien DNAs (viru ses and tr anspo sons). At least two differ-

ent types of silencing exist, one operating at the tran- cannot be overemphasized.

All of these factors have combined to make Arabi-scriptional and th e othe r at the p ost-transcriptional level

( Depicker an d Van Mon t ag u 1997). Elucidation of dopsis a premier organism with which to study problems

of intere st to all mo der n biologists. Many of these topicsthe mechan ism of silencing is ham pered by the inability

to direct a series of diagnostic constructs to the same are covered in this commemorative issue. The central


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475Perspectives

themes of modern biologydevelopment, transcrip- plant ( Waigmann an d Zambr yski 1995; Heinl ein et al.

1995). Perhaps cells that respond together for p lanttion, cell biologyall can be studied elegantlyin Arabi-

movemen t are united by an un derlying network of con-dopsis. Some of them, specificallythe response to gases,

nectivity via plasmod esmata. This speculation raisesare arguably studied best in Arabidopsis. Moreover, the

many questions. Are there cellular proteins, analogousplant-specific p roblems of p ho totropism, gravitropism,

to the viral movement proteins, that regulate the flowphotomorphogenesis, and disease resistance, formerlyof information through the plasmodesmata in responsestudied by classical gen etics or ph ysiological experi-to external signals? Are there transcellular cytoskeletalmen ts, are n ow being un raveled at the cellular level by

structures that extend through plasmodesmata fromthe application of genomics and the molecular geneticone cell to another? H ow do these hypothetical regula-techn iques discussed e arlier. Many of th ese findings aretory and cytoskeletal elements coordinate the cell walldirectly applicable to the improvement of crop s, whichexpansion that is thought to be n ecessary for motion?prob ably explains why this ined ible weed has sudden ly

If plasmodesmata provide the connections for con-become the darling of the agricultural schools.certed plant movements, then there must be some asAdvances in the genomics and molecular genetics ofyet u nd iscovered factor(s) that impose th e selectivityArabidopsis will perm it scientists to attack prob lems thatthat perm its one gro up of cells to respond to a stimulusformerlyseemed completelybeyond their grasp. I wouldand another to ignore it. Group identity could be con-like to ad dress two of these, the first theoretical and th eferred bya restricted pattern of intercellular connectiv-second practical.ity (only a subset of cells is connected by the samePlant movement: On e of the keyunan swered theoreti-plasmod esmal highway) and specific intercellular mo -cal qu estions in biology is: What is the basis for con-lecular signals. Th is possibilitycou ld be r evealed b y thecerted plant cell movements? We know that these dra-

analysis of mutan ts defective in response to th e stimu lusmatic movements are ind uced bye xternal stimuli (light,coupled with improved biochemical techniques for thegravity, and touch), but we do not have a conceptualidentification of plasmodesmal constituents. Once theframework that permits an understanding of the re-relevant molecules have been identified, new imagingsulting behaviors. This problem was addressed elo-technique s applied to wild-type an d movemen t mu tantsquently by Charles Darwin m ore th an a h und red yearsmight reveal a structural or chemical network sharedago:byth ose cells that respond in concert. Thu s, the solution

We believe that th ere is no structure in plants mor e won- to the problem of plant movement raised by Dar winderful, as far as its functions are concerned, than the tip

may not come from a unifying theoretical insight, butof the radicle. . . . It is hardly an exaggeration to say that

rather b y stitching togeth er the answers to many diversethe tip of the rad icle thus end owed, and having the p owerpuzzles.of directing the movements of the adjoining parts, acts

like the brain of one of the lower animals; the brain bein g The genetics o f nutrition: The remarkable develop-seated within the anterior en d of the body, receiving ments in Arabidopsis research will form the vanguard

impressions from the sense organs and directing the sev- of the third revolution in agriculture. As Jar ed Dia-eral movements.

mond has noted, Human historys most importantCha r l es Dar win, The Power of Movementevent since the last Ice Age was the rise of agriculturein Plants (1896)in southwest Asias Fertile Crescent ( Diamond 1997).

Dar win s comparison of the tip of the stem to the brain This first tsunami, which occurred in 9000 bc , was fol-of the lower animals focuses squarely on the issue, but lowed by another wave that occurred early in this cen-his metaphor is inapprop riate. Plants have no single tury: the engineering o f domesticated plants by the ap-organ, like the brain, that can be construed as directing plication of Mendelian geneticsin the 20s and 30s( seemovement. Moreover, movement in animals can be as- Figure 1). The third wave is building now, with thecribed to the position and con nectivityof the u nd erlying application of new DNA techn ologies. Already, in tro-anatomical structuresmuscles innervated by axons ductory botanical texts instruct todays college studen tslinked to th e centr al nervous system. But plant anatom y th at gen etic engine erin g will yield plant s th at make vita-has not r evealed the equ ivalent of a neural network that mins and drugs; resist drought, disease, and insects;could account for the concerted behavior of some cells and make better tasting food, clean the environment,but not others. recycle wastes, prevent tooth decay, and produce antibi-

Although plants do not have the equivalent of neural otics, biodegradable plastics, and fragrances ( Moo r enetworks, they have plasmod esmata, chann els thro ugh et al. 1998). A fewo f the m ore sober predictions alreadythe cellwalls that provide a continu ous cytoplasmic con- have been realized.nection between adjacent cells (Lucas 1995; Ding The scientific and social factors are clearlyin place for

1997). Plasmodesmata are portals for intercellular com- Arabidopsis to lead the third phase of the agricultural

mun ication as shown bytheir role in the spread of plant revolution, but the economic factor isnot. The difficulty

viruses; th ese viruses encode proteins that dilate th e isth at agricultural research is largelyfunded by the U.S.

Departmen t of Agriculture ( USDA), whose policies havechannels to enhance their movement throughout the


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476 G. R. Fink

year cryogenic immersion in the year 2173. The follow-

ing is the d iscussion between two nu tritionists observing

this modern Rip van Winkle:

Scientist 1: Has h e asked for an ything special?

Scientist 2: Yes, this m orn ing for bre akfast he re-

quested something called wheat germ,

organic honey, and tigers milk.

Scientist 1: Oh yes! Those were th e charmed sub-

stances that som e years ago were th ought

to contain life-preserving prop erties.

Scientist 2: You m ean there was no d eep fat, steak,

cream pies, hot fudge?

Scientist 1: Those were thought to be unh ealthy.

Precisely the opposite of what we now

know to be true.

Scientist 2: Incredible!

Figur e 1.The revolution in agriculture. The photographBut it is no joking matter. The prob lem with research

illustratesth e result of geneticen gineering in maize. Teosinte,in nutrition is that th ere is insufficient informationshown on th e left, is the wild grass thou ght to be the pro gentorabout th e two compon ents of the system: the p lants thatof modern corn. The first revolution involved domestication

and selection by natives of the new world. Th e second de- are the source of food and the animals that eat them.

pended upon Mendelian genetics to produce the cob on the The abilityto alter geneticallybo th parts of the equationright. In the center are the F1 progeny of a cross between would p rovide a powerful antidote to th e cur ren t chaos.teosinte and modern corn. The third revolution m ay produce

Although p lants are key to our d iet, we know little aboutplants that look d ramatically different from th ose of th e 20thhow they synth esize, metabolize, an d d eposit the n utri-century. The photograph was kindly provided by John Doe-

bl ey of the University of Minnesota. ents that are essential to our survival and longevity.

Much of this confusion could be remedied by using

Arabidopsis as a model for nutritional studies. At theremained conservative an d un imaginative in the face o f outset, it would be important to unravel the pathwaysthe momentous changes that could enable the agency by which fatty acids, vitamins, and other metabolitesto carry ou t its mission. Even with well-me anin g staff, are p roduced, how they are regulated, and how theirthe USDA has been unable to counteract the political deposition in the various tissues of the plant is con-forces th at historically h ave stymied its effectiven ess. Its trolled.largest competitive grant program, the National Re- On th e other side of the equation, there isa dearth of

search In itiative Com petitive Gran ts Program, has a woe- knowledge about howwe metabolize plant compounds.fullyinad equate bud get of only 97.2 million d ollars (FY Superimposed on this shortcoming is the problem that1998). Moreover, with an overhead rate of 14% it does the hum an population is geneticallyheterogeneous fornot pay the true costs of research, making its grants the ability to metabolize plant comp oun ds. Whats goodnon competitive with th ose from othe r federal agencies. to eat for one person may be life threatening for an-

One solution to this problem would be to place the other. The inability of people with phenylketonuria tofunding of plant engineering within the National Insti- metabolize phenylalanine is emblematic of thousandstutes of Health. The creation of a National Institute of of other genetically controlled metabolic differencesNutrition would house Arabidopsis in the appropriate in the human population. This genetic heterogeneitypantheon of Institutes dedicated to the health of the confounds current studies.nation. Such an act would not only provide support to It is worth contemplating how the field of n utritionaccelerate the revolution, it would br ing a breath of would be chan ged by genom ics and m olecular genetics.fresh air to nutrition, a field fundamental to public Think of the power of testing a batteryof mouse mutantshealth that has so far failed to ben efit from the ad vances consuming a diet of well-defined plant mutants. Suchin molecular genetics. Despite the enormous interest experiments would defin e the criticalnu trients that pro-in diet, there is little critical information about it; the mote good health, longevity, and resistance to infection.

professional journals and the popular pressabou nd with But, the benefits go much deeper. For example, one

contradictory claims bewildering to both scientists an d might find that a particular mouse mutant always dies

nonscientists. of coronary artery disease when fed a wild-type plant,

The confusion caused by the disarray in nutrition is but survives without complications when fed a mutant

captured poignantly in Woo dy Al l en s (1973) classic plant unable to produce a particular lipid. The ability

movie Sleeper, in which Miles Monroe, the owner of the to perform such experiments would turn nutrition into

an exact science.Happy Carrot Health Food Club, em erges from a 200-


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477Perspectives

One objection to this scenario is that Arabidopsis is LITERATURE CITED

not an ed ible plant. But, h ere th e spirit of the SecurityAl l en, W., 1973 Sleeper, United Artists.

Council should prevail: What we learn in Arabidopsis is Azpir oz-Leeh an , R., an d K. A. Fel dmann , 1997 T-DNA insertionmutagen esisin Arabidopsis: going backan d forth. Trend s Genet.directly applicableto the engineering of crop plan ts. Once we13: 152156.learn to make knockoutsand h omologous replacements

Bech t ol d , N., J. El l is an d G. Pel l et ier , 1993 In planta Agrobacte-in Arabidopsis these technologies will be quickly rium mediated gene transfer by infiltration of adult Arabidopsisadapted to oth er plants. Ther e seems to be no resistance thaliana plants. C.R. Acad. Sci. Paris 316: 11941199.

Bender , J., an d G. R. Fink, 1995 Epigenetic control of an endoge-to applying this idea to rapeseed, a close relative ofnous gene family is revealed by a novel blue fluorescent mutant

Arabidopsis that is a source of cooking oil. But, many of Arabidopsis. Cell 83: 725734.balk at the idea that Arabidopsis, a dicot, is a good Ch an g, S. C., S. K. Pa r , B. C. Kim, B. J. Kan g, D. U . Kim et al., 1994

Stable genetic transformation ofArabidopsis thaliana by Agrobac-model for the monocots (corn, rice, wheat, and sor-terium inoculation in planta. Plant J. 5: 551558.ghum), which constitute the major source of protein

Dar win, C., 1896 The Power of Movement in Plants. pp. 572573,for animals and humans. Appleton , New York.

However, the newfield of genomicshasprovided data Depicker , A., an d M.Van Mont agu , 1997 Post-transcriptional genesilencing in plants. Curr. Opin. Cell Biol. 9: 373382.suggesting that perhaps too much has been made of

Diamond, J., 1997 Location, location, location: the first farmers.the monocot/ dicot distinction ( Pat er son et al. 1996).Science 278: 12431244.

Although monocots and d icots are thought to h ave d i- Ding, B., 1997 Cell to cell transport of macromolecules throughplasmodesmata: a n ovel signaling path way in plants. Tre nds Cellverged from e ach oth er 130200m illion years ago, thereBiol. 7: 59.is considerable synteny between the two groups: large

Fl avel l , R. B., 1994 Inactivation of gene expression in p lants as atracts of genes in Sorghum are in the same order as consequence of specific sequence duplication. Proc. Natl. Acad.they are in Arabidopsis. This colinearity has direct conse- Sci. USA 91: 34903496.

Heh l , R., 1994 Transposon tagging in heterologous host plants.quences for the engineering of crop plants. Synteny Trends Genet. 10: 385386.between Arabidopsis and the monocots means that

Heinl ein, M., B. L. Epel , H. S. Padget t an d R. N. Beachy, 1995genes in Arabidopsis have functions similar to their or- Interaction of tobamovirus movement proteins with the plant

cytoskeleton. Science 270: 19831985.thologues in monocots, and knowing their position inKempin, S. S., S. J. Lil jegr en, L. M. Bl co k, S. D. Roun sl ey, M. F.Arabidopsis will aid in their isolation and manipulation

Yan of sky et al., 1997 Targeted disruption in Arabidopsis. Na-in crop plants. ture 389: 802803.

A second objection is that unlike mice, we are not Luca s, W. J., 1995 Plasmodesmata:intracellular channels for macro-molecular transport in plants. Curr. Opin. Cell Biol. 7: 673680.inbred and our gen etic diversity will preclude th e estab-

Mar t ienssen, R., 1996 Epigenetic phen omena: paramutation andlishmen t of a un iversally applicable diet. True en ough .

gene silencing in plants. Curr. Biol. 6: 810813.However, with the completion of the human genome Meyer owit z, E. M., an d C. R. Somer vil l e, 1994 Arabidopsis. Cold

Spring Harbo r Labo ratory Press, Cold Spring Harbo r, NY.project and the attendan t technologies for genotypingMoor e, R., W. D. Cl ar k an d D. S. Vod opich , 1998 Botany, p. 16,ind ividuals, it will be p ossible to iden tify the alleles asso-

McGra w-Hill, New Yor k.ciated with man y metabolic differences. On ce we und er- Pat er son , A. H ., T.-H. Lan, K. P. Reisch man n, C. Cha ng , Y.-R. Lin

et al., 1996 Toward a unified genetic map of higher plants,stand these human differences, it should be possible totranscending th e monocot-dicot divergence. Nature Genet. 14:design transgenic plants that better m eet ou r needs.380382.

The inclusion o f Arabidop sis in th e Security Coun cil Pr uit t , R. E., an d E. M. Meyer owit z , 1986 Characterizaton of theis noteworthy because it heralds a new era. Although genome of Arabidopsis thaliana. J. Mol. Biol. 187: 169183.

Redei, G. P., 1975 Arabidopsis as a genetic tool. Annu. Rev. Genet. 9:there will be many surprises along the way, on e thing111127.is clear: The emergence of a reliable and facile model

Sun da r esan, V., P. Spr ing er , T. Vol pe, S. Haw ar d, J. D. G. Jone splant that instructs both basic research and agriculture et al., 1995 Patterns of gene action in plant development re-

vealed by enhan cer trap and gene trap tran sposable elements.will alter the course of science and, ultimately, theGenes Dev. 9: 17971810.course of h istory. And I pred ict that when Sleepers Miles

Waigmann , E., an d P. Zambr yski , 1995 Tobacco mosaic virus move-Monroe awakens in the future, the nu tritionistswill want ment protein mediated protein transport between trichome cells.to know his genotype before he dines on the extraordi- Plant Cell 7: 20692079.nary plants of the 22nd century.

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