Cytological and Morphological Characterization of Anther Derived Plants from Sweet Pepper (Capsicum annuum L.) cv. "Special"

Surendra Lal Shrestha,Binod Prasad Luitel,Taek Jong Lee,Won Hee Kang 한국육종학회 2010 한국육종학회지 Vol.42 No.5

Anthers of sweet pepper F1 cultivar ‘Special’ were cultured on Dumas De Vaulx (C medium), supplemented with 0.1mgL-1 2, 4-D and 0.1 mg·L-1 kinetin with 3% sucrose, and 0.32% phytagel. The calluses obtained were further sub-cultured on Murashige and Skoog (MS) medium without growth regulators for regeneration. Regenerated plantlets were grown in plastic pots under plastic house and characterized their cytological and morphological characters in spring, 2008. Twenty percent plantlets were identified as haploid plants after chromosome and ploidy analysis. Haploid plants contained 12 chromosomes, high stomatal density with small stomatal length as compared to diploid plants. Stomatal length in haploids was 23.3% smaller than diploids. Haploid plants were characterized as small leaf and petiole size, poor vigor, thin stem and short plant height, short internodes and small flower buds, fruit size and fruit weight as compared to diploid plants and most of the haploid fruits were seedless. SP55, SP62, SP68, SP72 and SP77 are found high yielding double haploids with high total soluble content (8.6, 8.7, 9.2, 9.1 and 9.8 ºBrix, respectively) and desirable fruit shape, and recommended them to exploit as inbred lines for heterosis breeding.

Fruit Yield and Quality Evaluation of Sweet Pepper (Capsicum annuum L.) F1 Hybrids Derived from Inbred Lines

Surendra Lal Shrestha,Binod Prasad Luitel,Taek Jong Lee,Won Hee Kang 한국육종학회 2010 한국육종학회지 Vol.42 No.4

Sweet pepper inbred lines (KNU1003, KNU1006, KNU1007, KNU1009, KNU1015, KNU1017 and KNU2006) developed at Kangwon National University (KNU) through conventional means, inbred lines (5AVS1, 5AVS2, 5AVS3, 5AVS5, 5AVS7 and 5AVS8) collected at Rural Development Administration (RDA) and inbred lines (SP12, SP27 and SP14) derived from anther culture were used as female parents and anther culture derived homozygous lines (SP9, SP10, SP14, SP24, SP25, SP27, SP30, SP32, SP34, SP38, SP43, SP45 and SP51) were used as male parents to produce F1 hybrids. A total of 37 F1 hybrids were evaluated for fruit yield and quality characters in summer season, 2007. Variation in fruit number, fruit weight, fruit yield per plant and fruit volume was observed among the F1 hybrids. Superiority on yield over standard/commercial varieties were differed among F1 hybrids. Hybrid 5AVS8 x SP45 exhibited highest heterosis over Special (16.5%) and Fiesta (24.7%). Fruit quality characters (fruit length, fruit width, pericarp thickness, total soluble solid, fruit shape and fruit color) were varied among the F1 hybrids. Fruit number, fruit weight and fruit volume per plant were correlated with fruit yield. Based on the standard heterosis expressed by the hybrids and quality characters evaluation, KNU1017 x SP27, 5AVS1 x SP43, 5AVS5 x SP27, 5AVS8 x SP45, SP12 x SP38 and SP27 x SP25 hybrids were found to be superior over commercial cultivars and are selected. Inbred lines of these hybrid combinations can be used to produce F1 hybrid seed for commercial production.

Agro-morphological Characterization of Anther Derived Plants in Sweet Pepper (Capsicum annuum L. cv. Boogie)

Surendra Lal Shrestha,Binod Prasad Luitel,강원희 한국원예학회 2011 Horticulture, Environment, and Biotechnology Vol.52 No.2

This study was conducted in sweet pepper ‘Boogie’ to produce the haploids and diploids via anther culture, and to characterize their plant and fruit characters. Anthers were cultured on Dumas de Vaulx medium, (C-medium) supplemented with 0.1 mg・L-1 kinetin and 0.1 mg・L-1 2, 4-D, and 3% sucrose and 0.32% phytagel. Cultures were first incubated in dark at 35℃ for one week and then transferred at 25℃ under 16 h photoperiod for 40 days. Regenerated plantlets including haploids and diploids were evaluated for their plant and fruit characters at Hwacheon in spring, 2007. Results revealed that 39.6 % callus was formed in total anthers (6100) whereas only 1.78% plants were regenerated among them. Ploidy analysis showed that 78.1% regenerants were diploids. Haploid plants were characterized by short plant height, narrow leaf size, short internode length, small flower bud size, and small fruit with low volume than diploid plants. Average fruit volume measured in diploids was 62.7% higher than haploids. Variation in plant and fruit characters was observed within the diploid population and pepper breeder could utilize such variation in breeding program. Based on this characterization and compared to original cultivar, SP94, SP117, SP148, SP171 and SP174 are found superior diploid lines and recommended them to evaluate for variety improvement in sweet pepper.

Establishment of Hybrid Breeding System in Sweet Pepper

( Surendra Lal Shrestha ) 강원대학교 농업생명과학연구원(구 농업과학연구소) 2009 강원 농업생명환경연구 Vol.20 No.-

Sweet pepper (Capsicum annum L.) is a warm season crop that belongs to the solanaceae family. Itis a fast emerging vegetable and cash crop in Korea where seeds are quite expensive and most of the growing cultivars are exotic. Seventy percent growing area is under Kangwon Do. The main objective of this study was to create and select new genotypes of sweet pepper equal or better than existing hybrid cultivars. Anther culture was performed in Kangwon National University, horticulture laboratory and rests of the field experiment were performed in Hwacheon area. The anthers of widely grown hybrid cultivars (Derby, Debora, Fiesta, Special, Bossanova, Minipaprika, Clarity, Kufrah, Boogie, and Phenelene) were used in anther culture, where cultivar Kufrah had the highest percentage of callus formation (69.8%) but the percentage of plant regeneration were higher in Phenlene (2.67%) followed by Bossanova (2.41%). Even though there is the effect of genotype of donor plants on the success of anther culture, plantlets could be developed from all of these genotypes. These anther culture derived three hundred fourteen plantlets were morphological and cytological characterized. Twenty five percent plantlets were haploid which had shorter height and internodes, smaller leaves, plant structure, flower buds, fruit and stomata size, weak and poor in vigor and mostly seedless fruits with high total soluble solid contents as compare to diploid plants. Three plants were found male sterile. To determine the ploidy level of the plants obtained from in vitro anther culture, chromosome counts in root tip squashes and flow-cytometery were used. Distinct differences in stomata length was also noticed between haploid and diploid plants. One hundred twenty three inbred lines which were collected from abroad, inside the country and developed in the past were characterized on vegetative parameter (plant height, number of nodes to first branch, nodal color, maturity), flower and fruit characteristics (days to flowering, fruit set, fruit shape, fruit color, stalk cavity, apex grove), and yield parameter (average weight of fruit, yield per plant, and number of seeds per fruit). Variation in vegetative, fruit, and yield characters was observed. Seventeen lines were recorded with higher TSS content (>10.0%) where SP133, and SP91 had TSS 11.8%, and 11.7%respectively. Four lines (SP48, C01541, KNU3002, and SP142) had thicker pericarp (>6.0 mm).With respect to their over all performance, SP8, SP20, SP21, SP41, SP91, SP115, SP124, SP142, SP143, and KNU1009 were selected as superior inbred lines for cultivation.This characterization will help to identify the variety and to maintain purity in future. Beside this it will provide the information to the breeders on the varietal improvement work. Thirty eight female lines and twenty nine male lines were crossed and collected hybrid seeds of one hundred thirty combinations. These hybrid lines were grown in Vinyl house condition on field soil from last week of April to October in 2007 and 2008. Training and pruning were done by keeping three stems per plant. Nutrient and water were supplied through drip irrigation. The effect of crossing between male and female parents to the expression of characteristics in F1showed that; when small and big sized fruiting plants are crossed, F1 plants beard medium sized fruits, between red and yellow color all red color, between orange and yellow all yellow colored fruits, between round and lamuyo fruits it became glamour shaped fruits. Most of the hybrid lines were evaluated in 2007 and 2008. Among the hybrid lines thirty lines showed superior performance. In 2007, only hybrid cultivars; special and President, were grown as a control whereas in 2008, eleven commercial hybrid cultivars (Derby, Debla, Fiero, Special, Cupra, Fiesta, Jirisan, Maserati, Boogie, Plenty, and President) were grown for comparison. On the basis of good shape, taste, thick pericarp, high total soluble solid content, yield, and heterosis, seven hybrid lines; KNU1017(SP27), 5AVS5(SP27), 5AVS8(SP48), 5AVS7(SP45), 5AVS8(SP45), SP27(SP25) and SP12(SP38) were found superior, and selected for cultivation. These hybrid lines have heterosis on yield were 86%, 42%, 85%, 130%, 21%,67%, and 126%, respectively. Among them, 5AVS5(SP27) had yellow color fruit and the rest were red. In addition to this 5AVS1(SP43) had shown excellent performance as a minipaprika type containing high TSS, good taste, and conical shape. The other promising lines are KNU2006(SP14), KNU1006(SP14), KNU1009(SP27), KNU1015(SP32), 5AVS7(SP10), 5AVS8(SP32), 5AVS7(SP34), 5AVS7(SP27), 5AVS7(SP32), 5AVS8(SP10), 5AVS8(SP51), SP10(SP11), SP11(SP27), and KNU1015(SP27). The another experiment which was conducted to study whether priming with deep sea water results in enhancement of seed germination and to identify the optimum concentration of the priming solution, and duration of priming on sweet pepper (Cv. California wonder). Seeds were primed with five various concentrations (5%, 10%, 15%, 20% and 30%) for deep sea water for 48, 24, and 12 hours at 25℃. Priming in deep sea water (DSW) improved the early and final germination percentage, mean germination rate, emergence percentage and root and shoot length, compared with plain water, KNO3 and without priming treatments. 24 hours priming with 5 percentage DSW significantly improved the early germination percentage and radical length. It has also improved the mean germination and emergence days and early emergence percentage, compared with KNO3 and control. Hence the best seed priming treatment is 24 hours with 5 percentage DSW. Likewise in the study on the effect of deep sea water on root pruning of sweet pepper seedlings was done to minimize the excessive growth and root mat formation in Sweet pepper (Capsicum annuum L.) seedlings. Seeds were sown in plastic plug tray coated with 0%, 1%, 3%, 5%, 7%, 10% and 15% NaCl (w/v) and compared with control. Plants growth rate; shoot and root; length, fresh, and dry weight, and root structure were observed in seedling stage on plastic plug trays on 28, 35, and 42 days after sowing (DAS) respectively. Then same observation were recorded 35 DAT (days after transplanting). Results showed that NaCl coating had significantly minimized the excessive growth rate and root mat formation. Seedlings from the treatment produced more fibrous roots. Even though before transplanting, seedlings without treated were taller, longer root length, significantly higher fresh and dry weight of shoot and root but after transplanting, seedlings from NaCl coated trays showed better performance; taller plant, longer root, higher fresh and dry weight of shoot and root where 7% NaCl treatment gave the superior performance among the treatments and control. The optimum concentration of NaCl for coating plug tray appeared at 7% (w/v) to minimize root mat formation and to make appropriate growth.

Agronomic Variation in Anther Derived Plants of Sweet Pepper (Capsicum annuum L.) Genotypes

Shrestha, Surendra Lal,Luitel, Binod Prasad,Lee, Taek-Jong,Kang, Won-Hee The Plant Resources Society of Korea 2010 한국자원식물학회지 Vol.23 No.6

Anther derived double haploids (DHs) from sweet pepper genotypes ('Special', 'Derby', 'Bossanova', 'Fiesta', 'Debora' and 'Minipaprika') were used to study the agronomic variation in 2006. Ninety-nine successful DHs regenerants (32 from 'Special', 25 from 'Derby', 23 from 'Bossanova', 10 from 'Fiesta', 6 from 'Debora' and 3 from 'Minipaprika') were transplanted at plastic house and studied on their agronomic characters. Variation in agronomic characters was observed within the DHs of each genotype. DHs obtained from 'Derby' and 'Fiesta' exhibited wide variation in fruit yield $plant^{-1}$ whereas averaged fruit yield $plant^{-1}$ was highest in 'Derby' (1608 g) and less variation was observed in DHs of 'Bossanova'. Based on the agronomic characters expressed in DHs population at this environment, SP55, SP56, SP60, and SP116 from 'Special', SP8, SP10, SP14, SP16, and SP34 from 'Derby', SP115, SP119, SP142, SP143, SP196, and SP199 from 'Bossanova', SP41, SP45, and SP114 from 'Fiesta', SP21 from 'Debora' and SP91 from 'Minipaprika' identified as elite inbred lines and these DH lines could be used for commercial hybrids production in sweet pepper. Genetic relationship among the selected inbred lines using molecular markers and their response to diseases are further recommended to study.

Heterosis and Heterobeltiosis Studies in Sweet Pepper (Capsicum annuum L.)

Surendra Lal Shrestha,Binod Prasad Luitel,강원희 한국원예학회 2011 Horticulture, Environment, and Biotechnology Vol.52 No.3

Twenty-five sweet pepper inbred lines were used to study the heterotic response of economic traits at Hwacheon during 2006/07-2007/08. Out of 25 inbred lines, 12 inbred lines (KNU1006, KNU1015, KNU1017, KNU2006, CO1234, 5AVS1, 5AVS2, 5AVS7, 5SAVS8, SP12, SP27 and SP46) were used as female parents whereas 13 inbred lines (SP9, SP14, SP22, SP25, SP30, SP32, SP34, SP36, SP38, SP43, SP45, SP48 and SP51) were used as male parents and single crosses were made to produce F1 hybrid in 2006/07. Twenty-three F1 hybrids, their parents and three standard varieties (Special, Fiesta and President) were evaluated based on their fruit and fruit quality traits at plastic house in 2007. Variation of fruit yield and fruit characters was observed among the F1 hybrids, their parents and standard varieties. Hybrids of 5AVS7 × SP32, SP12 × SP38, 5AVS7 × SP45, CO1234 × SP32, KNU1015 × SP32, 5AVS7 × SP34, 5AVS8 × SP51 and SP27 × SP25 expressed the highest positive heterosis on fruit number and yield per plant whereas highest positive heterobeltiosis was exhibited by the cross 5AVS7 × SP32 (87.2%) and SP12 × SP38 (119.3%). Hybrid of 5AVS7 × SP32 exhibited the highest heterosis for fruit number (104.0%) and yield (141.2%) per plant. Hybrids of 5AVS7 × SP45, 5AVS7 × SP32, and 5AVS8 × SP48 had highest positive standard heterosis on fruit yield per plant over Special, Fiesta and President. Highest positive standard heterosis had manifested on fruit yield per plant using SP32, SP45, and SP48 inbred lines, which can be used as the male parents with their counterpart female for commercial hybrid production.

Effect of Genotype of Donor Plants on the Success of Anther Culture in Sweet Pepper (Capsicum annuum L.)

Surendra Lal Shrestha,강원희 한국자원식물학회 2009 한국자원식물학회지 Vol.22 No.6

To study the effect of genotype of donor plants on anther culture, anthers of nine hybrid cultivars (Derby, Special, Bossanova, Minipaprika, Fiesta, Boogie, Phenlene, Kufrah, and Clarity) of sweet pepper (Capsicum annuum L.) were cultured in a petridish containing C medium (Sibi, Dumas De Vaulx medium) supplemented with 0.1 mg/L 2,4-D and 0.1 mg/L kinetin, 3% sucrose and 0.32% Phytagel. The cultures were incubated in the dark at 35℃ for seven days, and then cultured at 25℃ with a photoperiod of 16 hr. daylength for 40 days. Frequency of callus formation and plant regeneration was varied among cultivars. Callus formation was ranged from 6% in Phenlene to 69.8% in Kufrah. The highest percentage of regenerated plantlets was obtained in cv. Phenlene (2.67%) followed by Bossanova (2.41%). Result of ploidy analysis; chromosome number observation and flowcytometry analysis, showed that haploid plants could be developed from all of these hybrid cultivars except cv. Fiesta, where highest percentage of haploid plants were obtained in Minipaprika (40%) followed by cv. Bossanova (36.1%). Haploid plants derived from these hybrid cultivars contained single set of chromosome (12 in numbers), higher stomata density (numbers), and smaller sized stomata as compare to diploid plants. The mean length of stomata was 26.9 ㎛ in haploid plants and 35.7 ㎛ in diploids.

Yield in Pepper (Capsicum annuum L.) Cultivars Established at Different Planting Bed Size and Growth Containers

Tumurbaatar Enkhsaikhan,Surendra Lal Shrestha,Won Hee Kang,Binod Prasad Luitel,Taek Jong Lee,Tserendendev Oyuntugs 한국원예학회 2010 한국원예학회 학술발표요지 Vol.2010 No.5

Effect of Deep Sea Water Seed Priming on the Growth of Rice (Oryza sativa L.) Seedlings

Yoon Byeong-Sung,Shrestha Surendra Lal,Kang Won-Hee The Plant Resources Society of Korea 2006 한국자원식물학회지 Vol.19 No.6

This experiment was conducted on rice (cv. 2005 Thaoi) seeds to study whether priming with deep sea water (DSW) results in enhancement of seed emergence and seedling growth and to identify the optimum concentration of Deep Sea Water (DSW) for priming. Two experiments were conducted subsequently. In experiment 1, four concentrations of the DSW (10%, 20%, 30% and 40%), and in experiment 2, five concentrations of DSW (10%, 15%, 20%, 25% and 30%) were prepared and seeds were primed for 24 hours duration at $25^{\circ}C$. Beside this, hydro priming with plain water was also included as a control. Experiments were laid out in Completely Randomized Design (CRD) with three replications. Result showed that 20% DSW seed priming treatment had improved the emergence, seedling height, number of roots and root length as compare to other with DSW or without DSW treatments. Beyond 20% DSW priming (i.e. 25%, 30% and 40%) were not suitable for priming the seed. On the basis of seedlings growth parameters; emergence, seedling height, root number and length, and shoot root ratio, 20% DSW priming was the best priming treatment.

The Effect of Deep Sea Water on Seed Priming of Sweet Pepper (Capsicum annum L.), Rice (Oryza sativa L.) and Ginseng (Panax ginseng C.A.Meyer)

Yoon Byeong-Sung,Shrestha Surendra Lal,Kang Won-Hee The Plant Resources Society of Korea 2006 한국자원식물학회지 Vol.19 No.3

This experiment was conducted to study whether priming with deep sea water results in enhancement of seed germination and to identify the optimum concentration of the priming solution, and duration of priming using sweet pepper (Cv. California wonder), rice (Cv. Ilpum) and ginseng seed. Sweet pepper and rice seeds were primed with 5 various concentrations (5%, 10%, 15%, 20% and 30%) for deep sea water for 48 hours, 24 hours and 12 hours at $25^{\circ}C$ and ginseng seeds in 5%, 10%, 15%, 20%, 25% and 30%, and 2,4,6, and 8 electrical conductivity (EC) which were made by desalinating deep sea water. Priming in deep sea water (DSW) improved the early and final germination percentage, mean germinal on rate, emergence percentage and root and shoot length, compared with plain water, $KNO_3$ and without priming treatments. In sweet pepper, 24 hours priming with 5 percentage DSW significantly improved the early germination percentage and radical length. It has also improved the mean germination and emergence days and early emergence percentage, compared with $KNO_3$ and control. Whereas, in rice, 48 hours priming with 10 percent DSW significantly improved the early germination percentage, plumule emergence percentage, root length and shoot height. Hence the best seed priming treatment on sweet pepper and Rice are 24 hours with 5 percentage DSW and 48 hours with 10 percentage DSW, respectively, whereas in ginseng, priming with EC4, EC8 and 25% DSW had shown better germination.