The present study was conducted in order to evaluate the existence and type of the dormancy and effects of different dormancy breaking treatments on seed germination of Colutea buhsei Boiss. (Fabaceae) as one of the Iran native plants, arid-land and ornamental shrub with ability of usage in the urban landscape design. The experiment was performed with 15 treatments and 3 replications in a completely randomized design. Seeds were subjected to different treatments including various levels of GA3, concentrated (98%) H2SO4, cold stratification (CS), soaking with tab water, floating in hot water (100�C) followed by continual cooling for 24 hr in the same water and combined treatments. Afterwards seeds were sown in laboratory conditions to determine the factors� effects on germination percentage (GP), germination rate (GR), root and shoot length of Colutea bohsei seeds. All of these treatments, except for GA3 (250 and 500 ppm), increased the percentage and rate of the seed germination. Maximum germination percentage (66.25%) and rate (14.9 seeds per day) in 7 days was obtained at concentrated (98%) H2SO4 (15 min). Maximum root and shoot length was observed at concentrated (98%) H2SO4 (15 min) plus GA3 (100 ppm, 24 hr). Use of GA3 (100 ppm) in 24 hr after H2SO4 increased the germination rate and shoot length but this additive effect was not significant

Ahmad BEIKMOHAMMADI

Ali TEHRANIFAR

Hossein NEMATI

Mahmoud SHOOR

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Available online at www.notulaebiologicae.roNot Sci Biol, 2012, 4(2):112-115Print ISSN 2067-3205; Electronic 2067-3264 Notulae Scientia BiologicaeStudy of Dormancy Type and Effect of Different Pre-Sowing Treatments on Seed Germination of Bladder-Senna (Colutea bohsei Boiss.) in Laboratory ConditionsAhmad BEIKMOHAMMADI*, Ali TEHRANIFAR, Hossein NEMATI, Mahmoud SHOORFerdowsi University of Mashhad, Agriculture Faculty, Horticultural Science Department, Azadi Square, Mashhad, Iran; ah.beik@yahoo.com (*corresponding author)AbstractThe present study was conducted in order to evaluate the existence and type of the dormancy and effects of different dormancy breaking treatments on seed germination of Colutea buhsei Boiss. (Fabaceae) as one of the Iran native plants, arid-land and ornamental shrub with ability of usage in the urban landscape design. The experiment was performed with 15 treatments and 3 replications in a completely randomized design. Seeds were subjected to different treatments including various levels of GA3, concentrated (98%) H2SO4, cold stratification (CS), soaking with tab water, floating in hot water (100°C) followed by continual cooling for 24 hr in the same water and combined treatments. Afterwards seeds were sown in laboratory conditions to determine the factors’ effects on germination percentage (GP), germination rate (GR), root and shoot length of Colutea bohsei seeds. All of these treatments, except for GA3 (250 and 500 ppm), increased the percentage and rate of the seed germination. Maximum germination percentage (66.25%) and rate (14.9 seeds per day) in 7 days was obtained at concentrated (98%) H2SO4 (15 min). Maximum root and shoot length was observed at concentrated (98%) H2SO4 (15 min) plus GA3 (100 ppm, 24 hr). Use of GA3 (100 ppm) in 24 hr after H2SO4 increased the germination rate and shoot length but this additive effect was not significant.Keywords: arid-land, drought tolerant, landscape design, native plant, ornamental shrubIntroductionThe Colutea L. (bladder-sennas) genus includes about 26 species of deciduous shrubs and small trees with a dis-tribution ranging from the Mediterranean region and South-Eastern Europe and North-West Africa (Browicz, 1963). Colutea bohsei Boiss. (Fabaceae) is a fast-growing shrub up to 3 m height, with flexible and gray stems and often grows at the altitudes of Alborz Mountains. The global distribution of this shrub is Iran and Turkmeni-stan; and its distribution in the Iran provinces is Golestan, Mazandaran, Azarbaijan, Khorasan, Semnan and Tehran. From botanical point of view, it bears branchy leaves com-posed of small and bright green leaflets, yellow flowers and fruit is a pod-shaped balloon (Mozaffarian, 2005). It is a drought-tolerant plant occurring in rocky and steep land-scapes and is known as an important species in preventing soil erosion and functional for landscape design. Colutea species are cultivated in temperate climates primarily for ornamental purposes (Krüsmann, 1984; Rudolf, 1974). Colutea L. species domestication is limited by the low ger-mination rates and high susceptibility of the seeds to fun-gal attack (Aguinagalde et al., 1990).Seeds of many woody plant species cannot germinate even if they are sown under optimal moisture, oxygen and soil conditions (Olmez et al., 2008). This problem is called dormancy and its causes are a hard and impermeable seed coat, immature or dormant embryo, absence of en-dosperm, or thick, fleshy seed cover (ISTA, 1993). There is great deal of variation in germination ability of seeds even within the same species. Poulsen (1996) reported that dormancy among and within seed lots, of the same species, varied with provenance, crop year, and individual trees.There are various germination obstacles in Colutea seeds (Dirr and Heuser, 1987; Dirr, 1990; Olmez et al., 2007, 2008) resulting in propagation difficulties (Heit, 1967; Piotto et al., 2003). There have been few studies to determine different methods and techniques to over-come seed dormancy in Colutea species. Generally, pre-treatments such as submersion in hot water, mechanical or chemical scarification, and hot aeration are used for seed coat dormancy while the cold and warm stratifications are usually applied to dormancy caused by restrictions at the embryo level (Landis et al., 1996). Among these methods and techniques, especially cold stratification, submersion in concentrated H2SO4, and steeping seeds in hot water (88-100°C), followed by 24 hr chilling, are well-known and used to increase germination percentage of Colutea seeds (Dirr and Heuser, 1987; Piotto et al., 2003; Olmez et al., 2007).The aim of this study was to investigate the dormancy type and influence of various dormancy-breaking treat-ments (submersion in tap water, submersion in hot water, submersion in H2SO4, GA3, cold stratification and com-Received 09 December 2011; accepted 07 April 2012Beikmohammadi A. et al. / Not Sci Biol, 2012, 4(2):112-115113GR= n1/t1 + n2/t2 + … + n15/t15;where n1, n2,..., n15 are the number of germinated seeds at times t1, t2,…, t15(in days).For the study of the treatments’ effect on root and shoot length, 15 normal seedlings were measured from each replication and expressed in centimeter in the final day of experiment. The experiment was conducted in ran-domized completely design with 15 treatments and three replications (60 seeds per replication) for each treatment. Data from the treatments were analyzed using SAS statis-tical software. The LSD test was used for treatment mean comparison (α = 0.05).Results and discussionExperiment analyse of variance indicates that effects of various treatments on germination percentage (GP), germination rate (GR), root and shoot length of Colutea bohsei seeds were significant (α = 0.01) (Tab. 1).Statistical analyses showed that the highest germination percentages (66.2%) were obtained in seeds submersed in H2SO4 for 15 min in laboratory conditions (Tab. 2). These results may indicate that the seeds of this shrub have a kind of dormancy that makes necessary their germination’s stimulation by removing the hard cover of seeds. Because the H2SO4 treatments are used in abrasion and softening the seed coat, it can be stated that the dormancy type of C. bohsei seeds is a physical dormancy and is pertaining to the seed coat. Similar findings were reported by Dirr (1990), who found that soaking Colutea seeds in H2SO4 for 30-60 min resulted in good germination. According to Dirr (1990), some Colutea seeds did not germinate easily unless the impermeable seed coat was ruptured by mechanical or chemical scarification. Olmez et al. (2008) stated that soaking C. armena seeds in H2SO4 for 30 min and sown in the greenhouse resulted in good GP value (77.2%). In addition, Olmez et al. (2007) reported that scarification of C. armena seeds by soaking in H2SO4 for 30 min and sown under laboratory conditions gave high GP (82.8%) and a short time to maximum GR (7 days). Ren and Tao (2004) stated that speed and percent germination of the Calligonum species can be greatly increased by mechanical scarification or acid treatments.The lowest average GP (7.5%) was determined at con-trol seeds and those floating in GA3 (250 and 500 ppm) for 24 hr, which were sown under laboratory conditions (Tab. 2). Dirr and Heuser (1987) reported that untreated Co-lutea seeds could be sown in autumn, but scarification was required for spring-sowing. Among combined treatments, bined this treatments) on germination of Colutea bohsei seeds, sown in laboratory conditions.Materials and methodsRipe fruits were collected in mid-summer, 2010 from altitudes (1500 m) of Shahrud (Kalpush) provenance with geographical co-ordinates: latitude 37°13’ North and lon-gitude 55°48’ East with relatively warm summers and cold winters located in the North part of Iran. The seeds were separated from the fruit and stored at 5±1°C in plastic bags in the refrigerator and then the following pre-sowing treatments were applied to determine their effects on ger-mination percentage (GP), germination rate (GR), root and shoot length of Colutea bohsei seeds:1-Floating in tap water (20±1°C) for 24 h;2-Floating in hot water (100°C) followed by continual cooling for 24 h, in the same water;3-Cold stratification (CS) for 15 and 30 days in (5±1°C) into refrigerator;4-Submersion in concentrated (98%) H2SO4 for 5, 15 and 30 min;5-Floating in GA3 (100, 250, and 500 ppm) for 24 h;6-Submersion in concentrated (98%) H2SO4 for 15 min, and then floating in GA3 (100, 250, and 500 ppm) for 24 h;7-Submersion in concentrated (98%) H2SO4 for 30 min, and then floating in GA3 (100 ppm) for 24 h;8-Control (no treatment).Seeds were stratified by putting layers of moistened sand and seeds on top of each other into refrigerator with mean temperature 5±1°C. Since there was a risk for some of the seeds to be mixed with the sand, because of their small size, linen cloth was placed between the sand and the seeds. The moisture of the sand and the seeds were checked continuously to avoid drying, heating and poor aeration. Twenty seeds were sown on wet filter paper in each glass petri-dish, under laboratory conditions (mean rela-tive humidity 50% and mean ambient day temperature of 23±2°C and 17±2°C night temperature) in Ferdowsi Uni-versity of Mashhad in the winter of 2011. Visible radicle growth was used to define germination. Seeds germination recorded daily; continued until no further germination oc-curred for 5 days; and recording duration of germination was 15 days. Germination percentage and germination rate was calculated using Maguire’s equations (1962):GP = 100(NG/NT)where NG is the number of germinated seeds and NT is the total number of seeds.Tab. 1. Summary of analysis of variance for measured parameters in Colutea buhsei Boiss. seedsMean squareSource of variation df Germination rate (GR) Germination percentage (GP) Root length Shoot lengthFactor 14 129** 1850** 1.73** 9**Error 45 4 58 0.05 0.06** Significant at p<0.01; df : degrees of freedomBeikmohammadi A. et al. / Not Sci Biol, 2012, 4(2):112-115114Tab. 2. Effect of various dormancy breaking treatments on germination percentage and germination rate of Colutea bohsei Boiss. seeds in laboratory conditionsTreatments Germination percentage (%)Germination rate(in day)Control (no treatment) 7.5 h 1.5 ghFloating in tap water (20 ±1°C) for 24 h, 22.5 f 4.1 fgFloating in hot water (100°C) followed by continual cooling for 24 h, in the same water, 18.7 fg 2.3 ghSubmersion in concentrated (98%) H2SO4 (5min) 38.7 dc 6 efSubmersion in concentrated (98%) H2SO4 (15 min) 66.2 a 14.9 abSubmersion in concentrated (98%) H2SO4 (30 min) 53.7 b 10.2 dFloating in GA3 (100 ppm) for 24 h, 30 de 8.7 deFloating in GA3 (250 ppm) for 24 h, 7.5 h 0.8 hFloating in GA3 (500 ppm) for 24 h, 7.5 h 0.4 hCold stratification (CS) for 15 days in (5±1°C) 11.2 gh 2.8 ghCold stratification (CS) for 30 days in (5±1°C) 21.2 fg 2.6 ghSubmersion in concentrated (98%) H2SO4 for 15 min + (GA 100 ppm) for 24 h 57.5 ab 13.3 bSubmersion in concentrated (98%) H2SO4 for 15 min + (GA 250 ppm) for 24 h 66.2 a 13.2 bcSubmersion in concentrated (98%) H2SO4 for 15 min + (GA 500 ppm) for 24 h 48.7 bc 10.4 cdSubmersion in concentrated (98%) H2SO4 for 30 min + (GA 100 ppm) for 24 h 66.2 a 17.4 aNote: Means in column with the same letter are not significantly different (LSD test, p<0.05)Fig. 2. Effect of combined dormancy breaking treatments on shoot and root length of Colutea bohsei seeds in laboratory conditions in the end of experimentFig. 1. Effects of various treatments on root and shoot length of Colutea bohsei seeds in laboratory conditions in the end of experimentBeikmohammadi A. et al. / Not Sci Biol, 2012, 4(2):112-115115and uses. Stipes Publishing Company, Champaign, IL.Dirr MA, Heuser CW (1987). The reference manual of woody plant propagation from seed to tissue culture. Varsity Pres, Athens.Heit CE (1967). Propagation from seed, successful propaga-tion of six hardseeded group species. Am Nurseryman 125(12):10-12.ISTA (1993). Rules for testing seeds. Seed Sci Technol 21:1-259.Krüsmann G (1984). Manual of cultivated broad-leaved trees and shrubs. Vol. 1, Timber Pres, Beaverton, USA.Landis TD, Barthell A, Loucks D (1996). Seed treatments to overcome dormancy. Forest Nursery Notes, United States, Department of Agriculture, Forest Services, USA, 9-12 p. Maguire JD (1962). Speed of germination-aid in selection and evaluation for seedling emergence and vigor. Crop Sci 2:176-177.Mozaffarian V (2005). Iran trees and shrubs, Farhang Maaser Press.Nadjafi F, Bannayan M, Tabrizi L, Rastgoo M (2006). Seed germination and dormancy breaking techniques for Ferula gummosa and Teucrium polium. J Arid Environ 64:542-547.Olmez Z, Gokturk A, Temel F (2007). Effects of cold stratification, H2SO4, submersion in hot and tap water pretreatments on germination of Bladder-Senna (Colutea armena Boiss. & Huet.) seeds. Seed Sci Technol 35(2):266-271.Olmez Z, Gokturk A, Karasah B, Yilmaz H (2008). Effects of cold stratification and H2SO4 pretreatments on germination of three provenances of smoke-tree (Cotinus coggygria Scop.) seeds in greenhouse and laboratory conditions. Afr J Biotechnol 8(19):4964-4968.Piotto B, Bartolini G, Bussotti F, Asensio A, García C, Chessa I, Ciccarese C, Ciccarese L, Crosti R, Cullum FJ, Noi AD, García P, Lambardi M, Lisci M, Lucci S, Melini S, Carlos J, Reinoso M, Murranca S, Nieddu G, Pacini E, Pagni G, Patumi M, Garcia FP,Piccini C, Rossetto M, Tranne G, Tylkowski T (2003). Fact sheets on the propagation of Mediterranean trees and shrubs from seed, 11-51 p. In: Piotto B, Noi AD (Eds.). Italy, Seed Propagation of Mediterranean Trees and Shrubs APAT, I.G.E.R. Poulsen K (1996). Case study: Neem (Azadirachta indica A. Juss.) Seed research, 14-22 p. In: Ouedraogos AS, Poulsen K, Stubsgaard F (Eds.). Proc an International Workshop on Improved Methods for Handling and Storage of Intermediate/Recalcitrant, Humlebaek, Denmark. Ren J, Tao L (2004). Effects of different pre-sowing seed treatments on germination of 10 Calligonum species. Forest Ecol Manage 195:291-300.Rudolf PO (1974). Colutea arborescens L. (Bladder-senna). In: Schopmeyer CS (Ed.). Seeds of Woody Plants in the United States, Agric. Handbook 450, USDA Forest Service, Washington DC, USA.submersion in concentrated (98%) H2SO4 for 30 min and then (GA3 100 ppm 24 hr) resulted the best germination percentage (66.2%) under laboratory conditions (Tab. 2). Similar results by Nadjafi et al. (2006), showed that acid scarification by H2SO4 (75% v/v), for 5 and 10 min with GA3 (1500 ppm, 48 hr), broke dormancy and induced 31.9% and 34.1% germination of T. polium, respectively. The best germination rates for laboratory conditions were 17.4 seeds in days which were determined in seeds soaked in concentrated (98%) H2SO4 for 30 min plus floating in GA3 (100 ppm) for 24 hr (Tab. 2).The highest root and shoot length was 3.6 and 4.9 cm respectively, which were observed in seed soaked (98%) H2SO4 for 15 min plus floating in GA3 (100 ppm) for 24 hr. Using of GA3 (100 ppm, 24 hr) after H2SO4, in-creased germination rate and shoot length but this addi-tive effect was not significant. In combined treatments (H2SO4+GA3) with increasing GA3 concentration, shoot and root length decreased (Fig. 2).ConclusionsAmong all the pre-sowing treatments applied, soaking in H2SO4 for 15 min resulted the highest GP (66.2%) and GR (14.9%) in the shortest time to maximum GR (8 days) with good root and shoot length that was 2.09 and 4.9 cm respectively. These results may indicate that the seeds of this shrub have a kind of dormancy that indicates their germination’s stimulation by removing the hard cover of seeds. Therefore, according to these results pre-treatment by submersion in H2SO4 for 15 min can be used to over-come dormancy of the C. bohsei Boiss. seeds.AcknowledgementsWe are thankful of financial support for Ferdowsi Uni-versity of Mashhad.ReferencesAguinagalde I, Perezgarcia F, Gonzalez A (1990). Flavonoids in seed coats of 2 Colutea species-ecophysiological aspects. J Basic Microbiol 30(8):547-553.Browicz K (1963). The genus Colutea L. monograph. Monographie Bot 14:136.Dirr MA (1990). Manual of woody landscape plants, their iden-tification, ornamental characteristics, culture, propagation Fig. 3. Seeds germination in laboratory conditions (A) and ma-ture shrub of  Colutea bohsei Boiss. in habitat (B)

Notulae Scientia Biologicae

Study of Dormancy Type and Effect of Different Pre-Sowing Treatments on Seed Germination of Bladder-Senna (Colutea bohsei Boiss.) in Laboratory Conditions

The present study was conducted in order to evaluate the existence and type of the dormancy and effects of different dormancy breaking treatments on seed germination of Colutea buhsei Boiss. (Fabaceae) as one of the Iran native plants, arid-land and ornamental shrub with ability of usage in the urban landscape design. The experiment was performed with 15 treatments and 3 replications in a completely randomized design. Seeds were subjected to different treatments including various levels of GA3, concentrated (98%) H2SO4, cold stratification (CS), soaking with tab water, floating in hot water (100C) followed by continual cooling for 24 hr in the same water and combined treatments. Afterwards seeds were sown in laboratory conditions to determine the factors effects on germination percentage (GP), germination rate (GR), root and shoot length of Colutea bohsei seeds. All of these treatments, except for GA3 (250 and 500 ppm), increased the percentage and rate of the seed germination. Maximum germination percentage (66.25%) and rate (14.9 seeds per day) in 7 days was obtained at concentrated (98%) H2SO4 (15 min). Maximum root and shoot length was observed at concentrated (98%) H2SO4 (15 min) plus GA3 (100 ppm, 24 hr). Use of GA3 (100 ppm) in 24 hr after H2SO4 increased the germination rate and shoot length but this additive effect was not significant

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Study of Dormancy Type and Effect of Different Pre-Sowing Treatments on Seed Germination of Bladder-Senna (Colutea bohsei Boiss.) in Laboratory Conditions

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