Characterization and Breakdown of Self-Incompatibility in Field ...

Characterization and Breakdown ofSelf-Incompatibility in Field Bindweed(Convolvulus arvensis L.)J. H. Westwood, T. Tominaga, and S. C. Weller

From the Horticulture Department, Purdue University,West Lafayette, Indiana (Westwood and Weller) and theF»culty of Agriculture, Shlnjhu University, Ina Nagano,399-45 Japan (Tominaga). This article Is Journal Paperno. 15175 of the Purdue Agricultural Experiment Sta-tion. Address correspondence to Dr. Westwood, Virgin-la Polytechnic Institute and State University, Depart-ment ol Plant Pathology, Physiology, and Weed Science,410 Price Hall, Blacksburg, VA 24061.

Journal of Heredity 1997^8:459-465; 0022-1503/97/S5.00

Field bindweed blotypes differ In glyphosate tolerance. Our previous studies haveindicated quantitative genetic inheritance of this trait. In order to study the geneticsof these differences we must be able to make sexual crosses, however, self-cross-ing and backcrossing are difficult due to self-incompatibility (SI) In field bindweedflowers. Therefore research was conducted to characterize and overcome the SIsystem In field bindweed. We determined SI to be of the muttlallelic, sporophytlctype based on observations of pollen-stigma interaction, and in the pattern of com-patibility of F, backcrosses to the parent biotypes. Pollen In incompatible crossesproduced short pollen tubes that failed to penetrate the stigma surface. Backcross-ing showed most F, plants to be compatible with one parent, indicating dominanceinteractions between alleles. SI In field bindweed was successfully overcome byboth bud pollination and heat treatment which exposed plants to 38°C for 2 daysbefore pollination, however, no seeds were produced. Treatment of stigma tips witha hot soldering iron for 2-3 s allowed abundant pollen tube growth from selfedpollen lying In a zone adjacent to the heat-injured area. This method resulted inproduction of viable selfed seed at rates equal to that observed in outcrosses.

Field bindweed is a twining, perennialweed that reproduces both by seeds andby adventitious shoots arising from aspreading root system. Because of its highreproductive capacity and competitivegrowth habit, it is a problem in crops suchas corn, small grains, sugar beets, andvineyards in all temperate regions of theworld (Holm et al. 1977; Weaver and Riley1982). Field bindweed is self-incompatible(Knight 1959) and thus an obligate out-crosser, which likely plays an importantrole in maintaining the high degree of phe-notypic variation observed in this species(Brown 1945; Call and Getty 1923; De-Gennaro and Weller 1984b; Garcia-Baudinand Darmency 1979; Kiss 1973; TominagaT and Weller SC, unpublished data). Thisvariation may be a factor in the inconsis-tent control of field bindweed by herbi-cides (Phillips 1978).

Self-incompatibility (SI) presents sever-al problems with respect to the study offield bindweed physiology and genetics.We are studying biotypes of field bind-weed to discover the physiological basisfor differential tolerance to glyphosate(DeGennaro and Weller 1984a). The gly-phosate tolerance trait was determined tobe heritable in an Initial dtallele study(Duncan and Weller 1987), but further ge-

netic analysis has been prevented becauseof SI. Selfing field bindweed would allowgenetic manipulations of this weed andwould facilitate physiological research inthat plants with homogeneous geneticbackgrounds could be studied.

Several mechanisms exist in plants forthe prevention of self-fertilization, andthese may be described according to theirgenetic control and physical manifestation(De Nettancourt 1977). In the broadestsense, SI may be classified as sporophyticor gametophytlc, based on how the pollenphenotype is determined. The pollen phe-notype in gametophytic SI is based on thegenetic complement of the haploid pollengrain, while the pollen phenotype In spo-rophytic SI is determined by that of thedlploid, pollen-producing plant. Sporo-phytic SI may be further categorized ashomomorphic or heteromorphic, depend-ing on whether all flowers are structurallysimilar or dissimilar. In homomorphic sys-tems, SI may be controlled genetically byone or more loci, each of which may havemultiple alleles. Also, SI systems may becharacterized by the site at which inhibi-tion occurs (i.e., stigma surface, style, orovary).

Self-incompatibility systems may be cir-cumvented to allow self-fertilization. Pol-

459Downloaded from https://academic.oup.com/jhered/article-abstract/88/6/459/840784by gueston 11 February 2018

llnatlon of stigmas in developing buds isamong the most common method, avoid-ing the SI response because the recogni-tion system is not fully developed until 1-2 days before anthesis (Nasrallah and Nas-rallah 1989). Other techniques have beenused to overcome SI, including treatmentof stigmas with an assortment of chemi-cals (growth regulators, amino acids, sug-ars), extracts from compatible pollen orother stigmas, UV irradiation, heat, elec-tric charge, and physical injury (Van Mar-rewijk 1989).

Self-incompatibility In the family Con-volvulaceae is homomorphic (De Nettan-court 1977), and although the SI system infield bindweed has not been studied, someresearch has been conducted on Ipomoeaspecies, which are also members of Con-volvulaceae. The SI system in /. setifera(Martin 1968), /. leucantha (Kowyama etal. 1980), and /. pes<aprae (Devall andThien 1992) was determined to be of thehomomorphic, sporophytic type, andsome researchers have successfully over-come SI in Ipomoea. Pollen tubes grew inself-crosses of /. Hstulosa after immersionof stigmas in 50°C water for 2 min (Prabhaet al. 1982). This disabled the SI system forat least 5 h, but it recovered by 24 h aftertreatment. Plant growth regulators (IAA,ethylene, and GA) also enhanced pollentube growth in self-crosses (Sood et al.1982; Kashyap and Gupta 1989a,b). Inthese studies pistils were excised to facil-itate treatment and observation of results.Despite successful promotion of self-crossed pollen tube growth, no selfedseed was produced.

Our objectives were to characterize theSI system In field bindweed and develop amethod for allowing selfed pollen tubegrowth and reliable production of self-crossed seed. This will allow further studyof the genetics of glyphosate response infield bindweed biotypes.

Materials and Methods

Four morphologically distinct biotypes offield bindweed collected from a single sitein Lafayette, Indiana, were used in ourstudies (DeGennaro and Weller 1984b). Allbiotypes (arbitrarily named 1, 2, 3, and 4)were self-incompatible, but produced seedwhen crossed with one another. Two ad-ditional biotypes, SD and H6, originatedfrom Saudi Arabia and were used for ex-periments on overcoming SI because theywere day neutral and flowered abundantlyyear-round. However, with respect to SI,these biotypes responded identically to

the Indiana biotypes. The original individ-ual plants were vegetatively propagated inorder to increase the number of floweringplants available for tests, but these cloneswere considered genetically identical andused interchangeably In crosses with oth-er biotypes and backcrosses with F,plants. The F, progeny of crosses betweenbiotypes were all labeled and treated asgenetically unique Individuals. Plants weremaintained in a greenhouse with supple-mental fluorescent and incandescent lighton a 16 h day, 8 h night regime to encour-age flowering. Plants were grown in a ster-ile soil mixture of topsoil, sphagnum peatmoss, and perlite (1:2:2 v/v) and surfacewatered as needed with a fertilizer solu-tion (200 ppm N and K, 60 ppm P).

Reciprocal crosses were performed byhand between all possible combinationsof the original four biotypes and the re-sulting F, seed were planted and grown tomaturity in the greenhouse. The F, plantswere named by a two-number designation,the first number representing the femaleparent and the second number the maleparent, that is, a cross using biotype 1 asfemale and pollen from biotype 3 was la-beled 13, and the reciprocal cross was 31.Several F, plants from each cross were ob-tained and each of these was backcrossedto its parents as both male and female todetermine its SI phenotype. The only ex-ceptions to this were the F, plants 23 and24 which did not flower under the givenconditions.

For SI determination studies, flowerswere emasculated and the stigmasbrushed with an anther from the male par-ent. Pollen tubes were allowed to grow forat least 4 h before harvesting the flowers.Stigmas were removed, squashed on aglass slide in a drop of aniline blue stain(0.1% aniline blue, 0.71% K3PO<), and pol-len tubes fluorescing bright yellow wereviewed with a microscope (Olympus BH-2)using UV illumination. The SI responsewas rated by the number of pollen tubesthat penetrated the stigma and grew downthe style according to the following scale:+ + = more than 15 pollen tubes, indica-tive of complete compatibility; + = 2-15pollen tubes, or moderate compatibility;and - = no pollen tubes, or complete in-compatibility. Control crosses using pollenfrom outcrosses were performed routinelyto confirm pollen viability. All SI responsesreported were verified by at least threereplications.

Bud PollinationsSelf-pollination of buds was performed onplants grown in the greenhouse as de-

scribed above. Pollinations were per-formed on buds up to 3 days before an-thesis, which required that petals be re-moved with forceps to expose the stigmaprior to pollination. Pollination was doneas described above, and the flower wasthen covered with a small envelope to pre-vent outcrossing.

General High-Temperature PollinationsGeneral high-temperature pollinationswere conducted after conditioning wholeplants in a growth chamber (Percival, witha Watlow microprocessor) for time peri-ods ranging from 0-5 days. Growing con-ditions consisted of 38°C, 16 h light (300p.E/m2/s), 8 h dark at 50% relative humid-ity. Pollen used for these studies was fromplants grown at ambient temperature Inthe greenhouse.

Localized High-TemperaturePollinationsLocalized, high-temperature stigma treat-ments were performed on flowers fromgreenhouse grown plants with a solderingiron (Radio Shack, 120 VAC, 60 Hz, 15 W/30W) set on 15 W and preheated for at least15 mln. Treatment consisted of gentlytouching the stigma tips with the iron for2-3 s, until slight withering of the stigmatips was visible. Within 5 min the injuredtip of the stigma would turn brown (Figure1). Pollen for self-pollinations was ob-tained from the treated plant and appliedwithin 10 min following heat treatmentalong the length of the stigma, to both In-jured and uninjured areas. For the experi-ment on duration of SI disruption, pollenwas applied at intervals between 30 s and24 h after treatment. However, since fieldbindweed flowers senesce after 1 day, the24 h pollination time required that flowersbe cut from the plant and held in water toavoid senescence of the stigma. For ex-periments requiring microscopic observa-tion of pollen tube growth, flowers wereleft on the plant for 4 h before harvest ofthe stigma for observation as describedabove.

Pollinations to obtain seed from self-crosses were performed as describedabove. Flowers were covered with an en-velope immediately after pollination toprevent outcrossing. Seed was allowed todevelop to maturity and harvested. Out-crossed controls were obtained by makingreciprocal crosses between the H6 and SDbiotypes. Seeds resulting from these vari-ous treatments were acid scarified by a 10min or 30 min immersion In concentratedsulfuric acid followed by a water rinse.

460 The Journal of Heredity 1997:88(6)

Downloaded from https://academic.oup.com/jhered/article-abstract/88/6/459/840784by gueston 11 February 2018

Figure 1. Field bindweed double-lobed stigma and style. Left: Uninjured stigmas. Right: Stigmas after solderingiron heat treatment applied to stigma tips. Magnification 15x.

Seeds were planted in sterile greenhousegrowth media (see above), placed in thegreenhouse, and observed for germina-tion.

Results

Characterization of the SI System inField BindweedSelf-incompatible crosses in field bind-weed were characterized by failure of pol-len tubes to penetrate the papillar cells ofthe stigma surface (Figure 2A). Most self-pollen grains did not produce pollentubes, though occasionally a short tube,no longer than a few diameters of the pol-len grain, would grow. These did not pen-etrate the papillar cells, but grew alongthe stigma surface. In compatible crosses,pollen tubes penetrated the stigma andgrew down the style within 4 h of pollina-tion.

Each of the field bindweed biotypes 1,2, 3, and 4 were crossed with each other,both as male and female. The F, seeds re-

Figure 2. Ruorescence micrographs of field bindweed stigma squashes. (A) Untreated stigma showing no pollen tube growth from selfed pollen. (B) High-temperature (solderingiron) treatment of stigma tip creates a zone of SI breakdown that allows growth of selfed pollen, p = pollen grain: pt = pollen tube. Magnification 50x.

Westwood et al • Self-Incompatibility in Reid Bindweed 461


Table 1. Compatibility of backcrowes between four Beld bindweed blotype* and their progeny Table 2. Effect of bad pollination forovercoming SI In field bindweed

F, generation(female)

12-131421313234414243

Paren-talblotype(fe-male)

1234

F, generation(male)

12

og

. .

13

0*0

Parental blotype(male)

1

%

033000*

17*

14 21

2

50

*33

0

0

*

31

3

plants compatible with parent blotype

*»0

000*

0

32 34 41 42

% of F, plants compatible with parent blotype

0100

* *60

25*0*

17100 * * 50

0 033 50 50

4

#60*

*0

335075

43

**

075

Days beforeanthests Self-cross Outcross

Pollen tube growth rating

0 - • + +1 - + +2 ++ + +3 ++ + +

Stigmas In developing buds of blotype SD were polli-nated with sell-or outcrossed pollen. Pollen tubegrowth was evaluated 4 h after pollination.

•Pollen tube growth rating scale: ++ = more than 15pollen tubes (complete compatibility), and - = no pol-len tubes (complete Incompatibility).

the H6 and SD biotypes. Furthermore, thenumbers of seeds produced per fruit afterselfing were 1.61 for H6 and 2.79 for SD,which were similar to those observed foroutcrossing these biotypes (Table 4).

Selfed seed was acid scarified and plant-ed to determine viability. The germinationrate of SD and H6 selfed-seed was 66% and

Values are the percent of F, plants from a given baclccross that were compatible with the parental blotype.' F, naming: Female biotype parent first, male blotype parent second* Indicates cross between nonrelated plants: high compatibility observed.

86%, respectively, while seed germinationof the outcrosses SD x H6 and H6 x SDwas 100% and 75%, respectively.

suiting from these crosses were thenplanted and the seedlings grown to matu-rity. F, plants were completely self-incom-patible when self-crossed (data notshown), but backcrosses between F,s andtheir parents were sometimes compatible,depending on the biotypes involved in thecross (Table 1). Generally F, plants wereincompatible with one or both parents(i.e., pollen from type 12 was compatiblewith stigmas of parental biotype 2 but notbiotype 1, and pollen from type 13 was notcompatible with either biotype 1 or 3).However, there were exceptions to thispattern, as some F, plants of the types 41and 42 were compatible with both parents.Variation in backcross compatibility wasalso observed between similar F, plantsfrom a given cross, for example, 50% of F,plants of type 12 were compatible with pa-rental biotype 2 while 50% were incompat-ible. With respect to reciprocal crosses,pollen from F, plant type 32 was compati-ble with biotype 2 stigmas in 100% of theF,s tested, but pollen from biotype 2 wasnever compatible with stigmas of type 32.

Overcoming SI In Field BindweedFor bud pollination studies with the H6and SD biotypes, flower buds were cross-and self-pollinated at 0, 1, 2, and 3 daysbefore anthesis. No pollen tube growth oc-curred on stigmas of open flowers or buds1 day before anthesis (Table 2). Pollen

tubes In self-crosses grew abundantlywhen pollen was applied to stigmas ofbuds 2 and 3 days before anthesis, how-ever, no seed was obtained. Outcrossedpollen grew abundantly at all time points.

General high-temperature treatment ofthe entire plant for time periods rangingfrom 0-5 days prior to pollination resultedin pollen tube growth in some self-crosses(data not shown). Maintaining plants for 2days or longer at 38°C resulted in a mod-erate level of selfed pollen tube growth.However, no seed was obtained from thesepollinated flowers. Outcrossed pollen pro-duced abundant pollen tube growth atthese temperatures, but also failed to pro-duce seed.

Localized high-temperature treatment ofstigmas resulted in both pollen tubegrowth and seed production in self-cross-es. Treatment of stigma tips with a solder-ing iron caused injury at the point of con-tact (Figure 1) and the selfed pollen pro-duced tubes only in the region of the stig-ma immediately adjacent the heat injury(Figure 2B). This disruption of SI was notreversible with time, as pollen tube pene-tration occurred up to 24 h after the high-temperature treatment (Table 3). Pollentube growth in this region was abundantand flowers produced seed in approxi-mately 70% of self-crosses (Table 4),which was equal to the seed producedfrom reciprocal outcrosses made between

DiscussionCharacterization of the SI System InField BindweedThe SI system in field bindweed is of themultiallellc, sporophytic type. Evidence tosupport this conclusion is present In re-sults from both the pollen-stigma interac-tion and the pattern of incompatibility inbackcrosses of F, plants to their parents.

The site of pollen tube inhibition is animportant feature of SI systems. In fieldbindweed, self-crossed pollen either failedto germinate or any pollen tube growthwas arrested immediately after germina-tion, with no stigma penetration (Figure2A). In general such pollen tube inhibition

Table 3. Duration of SI disruption followinglocalized high-temperature treatment of fieldbindweed stigmas

Time Intervalbetween heattreatment andpollination

30s60s10 mln30mln24 h

Pollen tubegrowth rating

+ +•+ ++ ++ ++

Stigmas of blotype H6 were heat treated with a solder-Ing Iron and then pollinated after various time Inter-vals.

•Pollen tube growth rating scale: ++ = more than 15pollen tubes (complete compatibility); + = 2-15 pollentubes.

4 6 2 The Journal of Heredity 1997.88(6)


Table 4. Effectiveness of localized heat treatment In prodaclng selfed seed In two blotypes of fieldbindweed

Percent of Dowers setting seed Number of seeds per fruit

Blotype

H6SD

Sell-cross

Control

4.54.3

Heat

70.066.7

Outcross

79.360.5

Seli-cross

Control

1.001.00

Heat

1.612.79

Outcross

1.562.65

Heat treatment consisted of contact with a hot soldering Iron, while controls were not heated. Blotype SD wasused as the source of outcross pollen for blotype H6 and H6 pollen was used for outcrosses with SD.

at the stigma surface is characteristic ofsporophytic SI, whereas pollen tubes inmost gametophytic SI systems grow intothe style before growth is stopped (Hes-lop-Harrison 1975). This difference be-tween systems was proposed to be due toimmediate recognition of self-pollen bythe interaction of a stigma papilla cell withsubstances in the pollen wall in sporo-phytic SI. However, exceptions to this ruleinclude the gametophytic SI systems ofOenothera and the family Gramlnae, whichalso show stigmatic inhibition (De Nettan-court 1977), so additional information isrequired to determine the SI system.

The existence of multiple biotypes thatare each self-incompatible, yet intercom-patible with one another, is consistentwith a multiallelic model of SI (presum-ably controlled by the S locus) (De Net-tancourt 1977). We assume that since fieldbindweed is self-incompatible, and thatthe plants used in this study were collect-ed from a wild population, the biotypesare all heterozygous at the 5 locus. Thusthe four biotypes used in the backcrossstudy may represent eight distinct 5 al-leles, though the current research cannotrule out the possibility of duplicate allelesor the influence of a second genetic locusinfluencing SI in field bindweed. Neverthe-less, with this assumption of 5 allele het-erogeneity, the backcross data support asporophytic SI system.

First, all field bindweed pollen respond-ed uniformly in backcrosses. In incompat-ible backcrosses, no pollen penetrated thestigma surface, whereas virtually all pollenin compatible backcrosses successfullyproduced pollen tubes (data not shown).This Indicates that all pollen share a sim-ilar SI phenotype and is consistent withsporophytic SI systems in which all pollentake the phenotype of the dominant S al-lele. In contrast, all backcrosses in a ga-metophytic SI system would be expectedto allow half the pollen to grow success-fully because only half the pollen wouldexpress the same S allele as the parent.

Second, the compatibility observed in

field bindweed backcrosses between cer-tain F, plants and their parents (Table 1)can be explained by S alleles acting indominance relationships, which is alsocharacteristic of sporophytic SI (Nasrallahand Nasrallah 1989). Where dominance re-lationships exist among 5 alleles, an F,plant is able to backcross to one of its par-ents because expression of one 5 allele ismasked by the other, more dominant 5 al-lele. Thus only the parent that donatedthe dominant allele will recognize pollenfrom the F, plant as self, while the parentthat donated the recessive allele wouldnot recognize the backcrossed pollen.Many of the F, field bindweed plants dis-played this compatibility with one parentbut not the other.

The fact that the backcross results werenot completely uniform may have to dowith both the heterozygosity of the parentplants and the fact that the dominance in-teractions of the 5 alleles may not followlinear hierarchies (Nasrallah and Nasral-lah 1989). Heterogeneity at the 5 locus ofthe parental biotypes means that sibling F,plants may differ from each other in theirS alleles, and thus all F,s would not giveuniform results in backcrosses. Indeed, inthe cases of backcross compatibility, usu-ally less than 100% of the F, progeny froma given cross are compatible. Neverthe-less, we can make some generalizationsabout 5 allele dominance in these fourfield bindweed plants. Biotype 3 was nevercompatible with any of its F, progeny, in-dicating that this biotype likely has one ormore very dominant S alleles. Biotype 4,on the other hand, was able to backcrosswith most of its F, progeny, so it probablycarries relatively weak 5 alleles, while bio-types 1 and 2 were intermediate betweenthese extremes. Further studies, includingselfing the parental biotypes to createplants that are homozygous at the 5 locus,will be required to fully characterize thedominance relationships between the S al-leles in these biotypes.

Variation in backcross compatibilitymay also be related to the variation in

dominance relationships depending onthe site of expression. In the sporophyticsystem of Brassica, dominance was foundto be more common In pollen, while in-dependent 5 allele action was more com-mon in stigmas (Nasrallah 1989; Thomp-son and Taylor 1966). This may explainwhy reciprocal crosses did not always be-have similarly, that is, F, plants of the type32 were compatible with only the blotype2 parent when used as male, but were in-compatible with both parents when usedas female.

Taken together the results presentedabove are entirely supportive of sporo-phytic SI in field bindweed, but are incon-sistent with gametophytic SI. Our resultsagree with work on SI in Ipomoea (Devalland Thien 1992; Kowyama 1980; Martin1968) and provide evidence from a secondgenus that SI in the family Convolvulaceaeis of the multiallelic, sporophytic type.

Overcoming SI in Field BindweedMany methods for overcoming SI havebeen explored over the last 60 years (VanMarrewijk 1989). These methods may in-volve either pollination before the SI sys-tem is active, or disabling an otherwisefunctional SI mechanism. The method ofchoice for overcoming SI depends on thetype of SI system and the species underinvestigation, although perhaps the mostwidely used method has been bud polli-nation.

Pollination of the bud stigma before theSI genes are fully expressed avoids the SIsystem. This method has been used in ob-taining self-crosses from the sporophytic SIsystem of Brassica (Nasrallah and Nasral-lah 1989) and the gametophytic system ofLycopersicon (Gradziel and Robinson 1989).In field bindweed self-crosses, pollen tubeswere produced that penetrated the stigmawhen applied 2 days before anthesis (Table2). Pollen tubes also grew at 3 days beforeanthesis, but at this time pollen tubegrowth was more disorganized than in pol-lination of mature stigmas. Pollen tubegrowth in buds was more random, withfewer tubes growing down the style. Theseresults indicate that the SI system in fieldbindweed becomes functional 1-2 days be-fore anthesis. This time frame for SI initia-tion is typical for sporophytic SI systems ofBrassica and /. Hstulosa where buds becomeself-incompatible 2 days before anthesis(Nasrallah and Nasrallah 1989; Prabha andGupta 1984). Self-pollination of field bind-weed buds was performed many times, butseeds were rarely obtained. This failurewas attributed to Immaturity of the repro-

Westwood et al • SelWncompatlbaity in Reid Bindweed 463


ductive organs and/or injury caused by re-moving sepals and petals. This is support-ed by results of bud pollinations in out-crossing experiments, where even compat-ible pollinations produced seed in less than20% of trials (data not shown).

High-temperature treatments have beenused to overcome SI in sporophytic sys-tems (Van Marrewijk 1989). Incubation ofBrassica flowers at 40°C for 15 min dis-rupted SI (Okazaki and Hinata 1987) as did50°C for 2 mln in /. fistulosa (Prabha et al.1982). Reid bindweed flowers incubated at50°C for 20 min resulted in self-pollen tubegrowth (data not shown). However, this isa very severe treatment, so milder tem-peratures were tested. A 2 day exposureof flowering plants to 38°C was required toobtain some tube growth of self-pollen(data not shown). The similarities of theheat period requirement and the time of SIonset based on bud pollination may notbe a coincidence. The relatively mild 38°Ctreatment may not disrupt an existing SImechanism, but may rather prevent prop-er development of the SI system. The spo-rophytic SI system of Brassica requirestwo gene products, a secreted glycopro-tein and a membrane-bound receptor kt-nase (Nasrallah and Nasrallah 1993). If asimilar mechanism is operating In Convol-vulus, high temperatures may upset theconfiguration of these proteins and lead toa failure in self-recognition.

Although self-pollinations of field bind-weed were performed under general high-temperature conditions, no seed was everobtained from these crosses. The hightemperature resulted in morphologicalchanges to plants that were held In thechamber for several days. New leaves andpetals decreased in size and thickened inresponse to the high-temperature stress,and perhaps seed production was also af-fected since outcrossed pollen did notlead to seed development under theseconditions.

Both bud pollination and heat treatmentrepresent deviations from normal pollina-tion conditions. Bud pollination relies onan immature ovary being receptive to fer-tilization, and heat treatment requires thatplants be held at stressful heat levels fordays. Although these conditions are con-ducive to overcoming SI, they are counter-productive to the ultimate goal of obtain-ing selfed seed. Therefore a method forheat-induced disruption of the SI systemwas tested that used mature flowers andcould be performed under greenhouseconditions.

Localized high-temperature treatment

using a soldering iron to heat stigma tipsof field bindweed resulted in self-pollentube growth. The intense heat produced aregion of damaged tissue at the stigma tip(Figure 1), but just below the Injured areawas a zone of papillar cells that allowedself-pollen to penetrate and grow (Figure2B). Outside of this zone, however, the SIsystem appeared intact and selfed pollendid not penetrate the stigma. The elongat-ed stigmas of field bindweed are ideal forthis treatment because the stigma tip canbe severely damaged by heat yet leave suf-ficient intact tissue at the stigma base forpollen tube penetration and growth. Rog-gen and Van Dljk (1976) first used a mod-ified, mini soldering iron to overcome SI inB. oleracea. They found that 70°C-80°C for2 s was sufficient to produce selfed seed,and in many cases was more effective thanbud pollination.

Unlike other heat treatments to over-come SI (Okazaki and Hinata 1987; Prabhaet al. 1982), SI did not recover in the stig-mas of field bindweed after localized heattreatment (Table 3). Selfed pollen tubeswere able to grow even 24 h after the heattreatment. Field bindweed flowers are onlyopen for the duration of 1 day, so for allpractical purposes the SI disruption waspermanent.

Localized high-temperature treatmentresulted in self-crossed seed production atrates equal to that of untreated outcrosses(Table 4). The variation between biotypesin number of seeds per fruit indicated thatnatural variation in seed production be-tween biotypes may be more importantthan pollen compatibility when the heattreatment was used. Perhaps a more im-portant factor In successful self-crossedseed production was the amount of pollendeposited in the narrow zone of SI disrup-tion, since only pollen in this relativelysmall zone will have a chance of fertilizingthe ovule. In the absence of heat treat-ment, about 4.5% of flowers producedseed, which is similar to rates reported for/. pes<aprae (3.4%; Devall and Thien 1992)and /. pandurata (8%; Stucky and Beck-mann 1982) when flowers were self-crossed without special treatment. Thisrepresents the level of natural breakdownin SI.

The self-crossing technique described inthis article is rapid, efficient, and can bedone under greenhouse conditions. Thefirst generation of self-crossed plants ger-minated well and appeared healthy. Theability to self-cross field bindweed willgreatly enhance our studies of the genet-ics and physiology of this important weed.

It is now possible to analyze the geneticsof glyphosate tolerance and to manipulatethe genetics of field bindweed plants formore in-depth study of other weedinesstraits.

ReferencesBrown EO, 1945. Notes on some variations In field bind-weed (Convolvulus arvensls L). Iowa St J Scl 20-.269-276.

Call LE, Getty RE, 1923. The eradication of bindweed.Agrlc Exp Sta Kansas St Agrlc Col Clrc 101:1-18.

DeGennaro FP, Weller SC, 1984a. Differential suscepti-bility of field bindweed (Convolvulus arvensls) biotypesto glyphosate. Weed Sci 32:472-476.

DeGennaro FP, Weller SC, 1984b. Growth and reproduc-tive characteristics of field bindweed (Convolvulus ar-vensis) biotypes. Weed Scl 32:525-528.

De Nettancourt D, 1977. Incompatibility In anglo-sperms. Monographs on theoretical and applied genet-ics, vol. 3. Berlin: Springer-Verlag.

Devall MS, Thlen LB, 1992. Self-incompatlbtllty In Ipo-moea pes<aprae (Convolvulaceae). Am Midi Nat 128:22-29.

Duncan CN, Weller SC, 1987. Herltabillty of glyphosatesusceptibility among biotypes of field bindweed. J He-red 78:257-260.

Garcla-Baudin JM, Darmency H, 1979. Differences ln-traspeclfiques chez Convolvulus awensts L Weed Res19:219-224.

Gradziel TM, Robinson RW, 1989. Breakdown of seli-Incompatlblllty during pistil development In Lycoperzi-con peruvianum by modified bud pollination. Sex PlantReprod 2:38-42.

Heslop-Harrlson J, 1975. Incompatibility and the pollen-stigma interaction. Annu Rev Plant Physlol 26:403-425.

Holm LG, Pancho JV, Herberger JP, and Plucknett DL,1977. The world's worst weeds. Honolulu: UniversityPress of Hawaii.

Kashyap R, Gupta SC, 1989a. A possible regulatory rolefor ethylene In the pollen-plstll Interaction In a sporo-phytic self-incompatibility system. Plant Growth Reg 8:127-135.

Kashyap R, Gupta SC, 1989b. The role of glbberelllcacid In the pollen-plstll Interaction In sporophytic self-lncompatlbillty systems. Plant Growth Reg 8:137-149

Kiss A, 1973. Morphological variations and herbicidesensitivity of Convolvulus awensts L. in the wine dis-trict of m6r. Acta Agron Acad Scl Hung 22:222-225.

Knight RJ Jr, 1959. Compatibility relations in the Con-volvulaceae In: Proceedings of the Ninth InternationalBotanical Congress, Montreal, Canada, August 19-29.Toronto: University of Toronto Press; 195.

Kowyama Y, Shimano N, Kawase T, 1980. Genetic anal-ysis of Incompatibility In the diplold Ipomoea speciesclosely related to the sweet potato. Theor Appl Genet58:149-155.

Martin FW, 1968. The system of self-incompatibility InIpomoea. J Hered 59:263-267.

Nasrallah JB, Nasrallah ME, 1989. The molecular genet-ics of self-lncompatlbllity In Brassica. Annu Rev Genet23:121-139.

Nasrallah JB, Nasrallah ME, 1993. Pollen-stigma signal-Ing In the sporophytic self-incompatibility response.Plant Cell 5:1325-1335.

Nasrallah ME, 1989. The genetics of self-incompatibilityreactions in Brassica and the effects of suppressergenes. In: Plant reproduction: from floral Induction topollination (Lord E, Bemler G, eds). American Societyof Plant Physiologists Symposium Series, vol. 1. Rock-vllle: Society of Plant Physiologists; 146-155.

4 6 4 The Journal of Heredity 199788(6)


Okazaki K, Hlnata K, 1987. Repressing the expression Roggen G, Van Dl)k AJ, 1976. "Thermally aided polll- Thompson KF, Taylor JP, 1966. Non-Unear dominanceof self-Incompatibility In cruclfers by short term high nation": a new method o( breaking self-lncompaUbillty relationships between S alleles. Heredity 21:345-362.temperature treatment. Theor Appl Genet 73:496-500. In Brassica oleracea L Euphytlca 25:643-646. Van Marrewljk GAM, 1989. Overcoming Incompatibility.Phillips WM, 1978. Field bindweed: the weed and the c . „ ^ L L ^ In: Manipulation ol fruiting (Wright CJ, ed). London:problem. Proc N Cent Weed Cont Conf 33:140-141. S ^ *• P™*503 *. Govi' S. Gupta S, 1982. Overcoming Butterwcrths; 173-191.

Prabha K, Sood R, Gupta SC. 1982. High temperature- i S m S T * "" "Xm°e° *""*" * ^ ' " ^ ^ I f v ^ T ^ ^ "I?"*? p°/Induced inactivation of sporophytic selHncompaUblll- " -1 1-3 3 3-3 3 9- ^ Co"UOiVU'US aWemtS L Om J P l a n t

22p p y

ty In Ipomoea Kstulosa. New Phytol 92:115-122. Stucky JM, Beckmann RL, 1982. Pollination biology, sell-Prabha K, Gupta SC, 1984. Effects of floral age and flow- incompatibility, and sterility In Ipomoea pandurata ( L ) ^ ^ t i P i L , ^ oocerlng season on seU-lncompaUbUlty vigour In Ipomoea & F. W. Meyer (Convorvulaceae). Am J Bot 69:1022- Accepted November 29, 1996fistulosa. Beltr Blol Pflanzen 59:359-366. 1031. Corresponding Editor Prem P. Jauhar

Westwood et a) • Self-Incompatibility in field Bindweed 4 6 5


Characterization and Breakdown of Self-Incompatibility in Field ...

Documents

Transcript of Characterization and Breakdown of Self-Incompatibility in Field ...