Prec. Indian Acad. Sci., Vol. 87 B, (Plant Sciences-2), No. 5, May 1978, pp. 129-133, ~) Printed in India.

Effect of x-irradiation on physiological and morphological variability in Abelmoschus esculentus (L.) Moench

SRINATH RAO and DIGAMBE R RAO Department of Botany, Kakatiya University, Vidyaranyapuri, Warangal 506 009

MS received 4 October 1977; revised 4 March 1978

Abstract. The variations induced by x-irradiation of seeds of Abelmoschus esculentus in xz generation have been studied. Percentage of germination showed a slight in- crease in treatments at 1 to 3 kR, while at the higher doses there was a gradual de- crease in germination. Seedling growth measured at regular intervals of 6 hr up to 96 hr, showed that growth rate was reduced and germination delayed at doses of 8 to 10 kR. Lateral root number and length were not significantly effected by the dosage rate. Total chlorophyll content of leaves was reduced by 50~o at 8 kR treat- ment, while an increased chlorophyll metabolism was observed at doses of 4 and 7 kR. Increasing dose rate caused a gradual decrease in plant height. Early flower- ing was induced in 8, 9 and 10 kR treatments, while flowering was delayed in the 5 and 6 kR treated samples. Average pod length did not show significant variations between the different dose rates as was also the stomatal index. Increased seed weight was observed in samples treated above 8 kR.

Keywords. x-irradiation; percent germination; seedling growth; chlorophyll meta- bolism; plant height; Okra.

1. Introduction

Abelmoschus esculentus Moench (=Hibiscus esculentus Linn.), a member of the family Malvaceae, is the most commonly cultivated vegetable crop (Okra) through- out India. Apart from its value as a vegetable, the capsules and seeds of the plant have been reported to have medicinal value (Chopra et al 1956, 1969). Since the time of Muller (1927) and Stadler (1930), several workers have investigated induced mutagenesis in plants and animals through irradiation by x-rays (Johnson 1931; Sax and Swanson 1941; Rawlings et al 1958; Gladstone and Hunter 1958; Cardona et al 1960). Interspecific hybridization followed by ovule and embryo culture tech- niques has been successfully tried by Gadwal et al (1968).

Radiation-induced mutagenesis in Okra was initiated by Abdel-A1 (1966) by using gamma rays. Apart from the work of Durra (1969), mutagenic studies on the plant have been scanty. Nandpuri et al (1971) studied the radiation-induced variability in the plant through gamma irradiation. Gupta and Yashvir (1975), and Yashvir (1975, 1977) investigated the mutagenic effects of individual and combined treat- ments of gamma rays and ethyl methane sulphonate (EMS) in this species.

The present paper embodies the results o f investigations on the effect of x-irradia- tion of dry seeds o f Okra on the germination, chlorophyll metabolism, etc. in the x 1 generation.

129

130 Srinath Rat and Digamber Rat

2. Materials and methods

Seeds of the cultiver Pusa Sawani obtained from the National Seeds Corporation of India, Warangal Branch, were subjected to x-irradiation at the Govelnment Mahatma Gandhi Memorial Hospital, Warangal. The doses given were 1 to 10 kR at the increasing dose rate of 1 kR. For each treatment including the control (untreated) 200 seeds were used. Seeds were sown in a field for randomized blocks with three replicates fifty seeds in each replication, per treatment. Seeds were sown one foot apart in the row, with each replicate consisting of five rows. The remaining 50 seeds in each treatment were soaked in tap water for 6 hr, and then transferred to sterilized petri dishes lined with moist Whatmann No. 1 filter papers. Five replicates with ten seeds in a petri dish for each treatment were maintained in order to study the effect of different x-ray doses on the germination and growth of the seedlings. Stomatal index was calculated according to Salisbury's (1928) method. For the estimation of total chlorophyll content in the leaf, Amon's (1949) procedure was adopted. Care was taken to select leaf material at random and of the same age from different plants. Seedling measurements were made from the germinating seeds in petri dishes, kept at a temperature of 21-4-2~

3. Results and discussion

3.1. Per cent germination

The lower doses (1 to 3 kR) of x-rays showed a slight increase in the germination percentage of seeds (75"5 to 78.4 70) over that of the control samples (74-6 7o). How- ever, an increased dosage rate of 4 to 10 kR caused a gradual decline in the per cent germination of seeds (73.1 to 57"5 70). Nandpuri et al (1971) observed that high exposures to gamma rays increased the mortality of seeds in this species. It would seem that x-rays are somewhat milder in its effects as compared to the gamma irradiation, at 10 kR exposure, the x-ray exposed seeds showed 57.6 700 germination (present study), while the gamma irradiated seeds showed only 40-5 700 germination (Nandpuri et al 1971). According to Sax and Swanson (1941) radiations effect aberrations in chromosomes of the individual cells but in tissue reaction to the radiations, physiological effects are more important. Cardona et al (1960) reported decreased germination of kidney beans exposed to high doses of x-rays. Gladstone and Hunter (1958) observed that in tomato seeds sensitivity to x-rays increased ten fold as the water content of seeds decreased from 12"2 to 1-2 70- Such a change in sensitivity to x-rays might be of relevance in this species also.

3.2. Seedling growth

Seedling growth was determined by taking measurements at 6 hr intervals for 96 hr and the final measurements were made on one week-old seedlings. The data pre- sented in table 1 show that higher doses (8 to 10 kR) caused a reduction in the growth rate of the seedlings. Germination was delayed occurring only 42 hr after soaking, in contrast to the controls in which germination was observed in 36 hr. The lower doses (I to 4 kR and 7 kR) showed precocious germination, even over the controls,

X-irradiation effects in A. esculentus

Table 1. Effect of x-rays on seedling growth at different intervals.

131

Duration kR (in cm) in hr 0 1 2 3 4 5 6 7 8 9 10

6 . . . . . . . . . . . 12 . . . . . . . . . . . 18 - - 0-10 0"10 0"10 0"10 - - - - 0 .10 - - - - - - 24 - - 0"30 0-20 0'50 0"40 0"10 - - 0"50 - - - - - -

30 - - 1"30 0"50 0"80 0"90 0"40 0'10 0.90 - - - - - - 36 0"50 2"00 1.00 1"30 1.00 0"90 0"40 1"50 - - - - - - 42 0.90 2"90 1.50 1"90 1"80 1.10 0"70 2"10 - - - - 48 1"30 3.30 2.00 2"40 2"30 1.30 1"20 2"70 0'10 0"20 0"20

54 1"80 4.00 2"60 3"00 2.90 2"40 1"90 3"80 0"50 0"50 0.50 60 2"60 4"80 2.90 3"60 3.40 2.60 2"40 4-10 1"10 1"10 0-96 66 2"90 5"50 3 " 3 6 4"10 3.90 3"10 3"10 5.70 1"80 1"70 1"60 72 3.00 6.00 3"70 4"80 4"30 3"70 3"70 6"30 2"41 2"10 2"10

78 3.20 6"52 4.00 5"40 4"90 4"10 4"30 7.00 3.00 2 " 4 6 2.46 84 3.80 7"00 4"10 5'90 5.50 4"40 4"50 7.50 3.50 3"10 3"10 90 4-30 7"30 4"50 6-20 6.10 4"80 5.00 8"00 3"75 3"50 3.91 96 4.60 7"80 4"90 6"70 6"70 5"20 5"30 8.40 3"95 3"85 4.00

Table 2. Effect of x-rays on length, number of lateral roots and length of lateral roots in one week old seedlings.

D o s e Length of seedlings Number of lateral Length of lateral roots rate in cms roots per seedling per seedling in m m

(in kR) Range Mean S.E. C.V. Mean S.E. C.V. Mean S.E. C.V.

0 9.2-14"5 10"90 2 " 1 9 20"09 12.00 2"30 19"16 8.00 1"12 14"00 1 9.0-14"7 11"05 2"36 21.36 15.00 2"68 17"36 14"50 2"89 19"93 2 9.0-14-7 11.10 1.93 1 7 " 3 9 12.40 2"28 18"33 12-10 2-42 20.00 3 10"2-14"3 11.51 2"16 1 9 " 3 7 12"40 2"19 17"66 12"62 1"68 13"33

4 14.9-20"7 16.98 1-66 9"78 12.30 1"81 14"71 12"32 1"45 11"77 5 15"0-24-0 18.00 2"30 1 2 " 7 7 13.40 2"12 15"82 10"50 2"71 25.81 6 11.1-15"4 13.16 2"61 19"83 12.00 1"45 12"08 10"28 2'31 20.72

7 15.1-20"7 17.04 2"41 1 4 " 1 4 11"30 2"66 23"53 13.60 2"30 16"91 8 9.3-10"8 10.10 1"53 1 5 " 1 5 13.40 1"89 14"10 10.20 1"88 18"43 9 8.0-13-0 10.90 2-03 1 9 " 6 2 1 3 " 0 8 2"26 17"38 11.08 2"04 18"41

10 8-5-11"8 10.90 1"82 1 6 " 6 9 12.30 1"11 9"08 10.62 2"50 23"54

occurring at 18 hr after soaking. Seedling length taken at the end of one week showed that the best growth (18 cm) occurred in 5 kR treatments whereas the lowest growth (10"9 cm) was in 9 and 10 kR treatments (table 2).

3.3. Lateral root number and length

The counts made on one week-old seedlings (table 2) showed that increased lateral rooting was induced in 8 and 9 kR treatments. The lateral roots attained an aver- age maximum length of 14.5 mm in 1 kR treatment. The control samples and the other treatments showed a varying mean length of 8 to 13"6 ram. In this respect the dose rate does not seem to have proportionate effect on lateral root number or

length.

132 Srinath Rao and Digamber Rao

Table 3. Effect of x-rays on height of the plants (50 days after sowing) and chlorophyll content.

Dose Height of the plants in eras Chlorophyll rate content in (in kR) Range Mean $.E. C . V . rag/gram wt

0 36"0-51"2 44.45 2"32 5"22 0-26 1 30.5-50.2 40"26 2'31 6.98 0.26 2 30"2-49"3 39"96 1"51 3-78 0.20 3 29"5-50"1 39"36 2"48 6.22 0"33 4 29.3-47.7 37.30 1"30 3"49 0.54 5 30.3-42"4 36"66 2'32 6"33 0.43 6 29"3-46"3 35.97 2"15 5.98 0.33 7 27.5-40.3 34.55 3.94 11.40 0.56 8 28.4-36.5 33.04 2.20 6.66 o. 11 9 26.6-33.5 31.10 2.53 8.14 0.48

10 28.7-35.3 30.71 1.84 5.99 0.17

3.4. Effect on chlorophyll metabolism

Effect of x-rays on chlorophyll metabolism was measured in terms of mg of chloro- phyll per gram wt of leaf. Data presented in table 3 show that 4 kR and 7 kR treat- ments caused an increase in the chlorophyll content of leaves, while a dose of 8 kR resulted in more than 50 ~o reduction of the content over the control sample. Gupta and Yashvir (1974-75), and Yashvir (1975) have reported the development of chlo- rophyll and non-chlorophyll chimeras in this plant species subjected to different doses of gamma rays.

3.5. Plant height

Height of the plants was measured 50 days after sowing. Increasing dose rate, a gradual suppression of plant height was observed (table 3). Gupta and Yashvir (1974-75) reported that treatment with EMS for a longer duration caused a de- crease in the seedling height in this species; however, when EMS (1 ~) and gamma rays (40 kR) were combined, the effect of EMS was reduced. Yashvir (1975) work- ing on this species, reported that greater reduction in mean value of plant height was obtained by gamma ray treatment.

3.6. Flowering

Early flowering was induced in 8, 9, and 10 kR treatments. The plants flowered in 42 days while lower doses (5 and 6 kR treatments) caused late flowering requiring about 50-51 days for induction. On the other hand gamma-ray treatment of the species has shown that a dose of 60 kR caused early flowering but treatment with half the dose rate caused delayed flowering (Nandpuri et al 1971).

3.7. Fruits and seeds

There was no change in fruit colour in any of the treatments. Average pod length did not show any significant variations between the various treatments although a

X.irradiation effects in A. esculentus 133

pe rcep t ib l e reduc t ion was discernible wi th increasing doses. G a m m a rad ia t ion has g iven s imi lar resul ts (Nandpur i et al 1971). Average weight o f 25 a i r dr ied seeds was f o u n d to be m a x i m u m in 8 k R and m i n i m u m in 9 k R t rea tments .

3.8. Stomata l index

S t o m a t a l index values der ived for the lower epidermis o f leaves o f the same age g roup d id n o t show significant va r ia t ion in the mean values. The mean index va lue however , was m a x i m u m in 7 k R t rea tment , and m i n i m u m in 8 k R t rea tment .

Acknowledgements

The au thors are gra teful to Prof. U B S Swami for facilities, and to Prof. K Venka ta Ramiah , for encouragement . They are also thankfu l to D r G Venkat Reddy, Gove rnmen t M G M Hospi ta l , W a r a n g a l for p rov id ing i r rad ia t ion facilities. One o f us (DR) is deeply i ndeb ted to Prof. Ja fa r N i z a m a n d Prof . V S R a m a Das for ini t ia t ing h im in this l ine o f work.

References

Abdel-Al Z 1966 Studies on the possibilities of using gamma radiations to induce mutations in Okra (Hibiscus esculentus L.); Alexandria J. ,4gri. Res. 14 283-290

Amen D I 1949 Copper enzymes in isolated chlorplasts. Polyphenol oxidase in Beta vulgaris; Plant Physiol. 24 1-15

Cardona C, Daurte R and Mansini S 1960 The use of artificial radiation in agriculture and its appli- cation to kidney been in Colombia; Agri. Trop. 16 514-523

Chopra R N, Nayar S L and Chopra I C 1956 Glossary o f Indian Medicinal plants (New Delhi: CSIR) p. 133

Chopra R N, Chopra I C and Varma B S 1969 Supplement to Glossary o f Indian Medicinal Plants. (New Delhi: CSIR) p. 38

Dutta O P 1969 Breeding Okra for quality, yield and resistance to virus and insect pests; Half yearly report Inst. Hort. Res. Bangalore

Gadwal V R, Joshi A B and Iyer R D 1968 Interspecific hybrids in Abelmoschus through ovule and embryo culture; Indian J. Genet. 28 269-274

Gladstone J S and Hunter A W S 1958 Effects of seed moisture storage on the growth and survival of xx tomato plants; Can. J. Genet. Cyt. 1 339-346

Gupta P K and Yashvir 1974-75 Mutagenic effects of individual and combined treatments of gamma rays and EMS in Okra (Abelmoschus esculentus Moench.); J. Cytol. Genet. 9 & 10 93-97

Johnson Edna L 1931 On the alleged stimulating action of x-rays upon plants; Am. J. Bet. 18 603- 614

Muller H J 1927 Artificial trans-mutation of the gene; Science 66 84-87 Nandpuri K S, Sandhu K S and Randhawa K S 1971 Effect of irradiation on variability in Okra

(Abelmoschus esculentus (L.) Moench); J. Res. 8 183-188 Rawlings I O, Hanway D G and Gardner C O 1958 Variations in quantitative characters of soybean;

Agron. J. 50 524-528 Salisbury E J 1928 On the causes and ecological significance of stomatal frequency with special

reference to Woodland flora; Phil. Trans. R. Soc. London B216 1-65 Sax K and Swanson C P 1941 Differential sensitivity of cells to x-rays; Am. J. Bet. 28 52-59 Stadler L J 1930 Some genetic effects of x-rays in plants; J. Hered. 30 1-19 Yashvir 1975 Induced quantitative mutations in Okra (Abelmoschus esculentus Moench). I. Plant

height mutations; Prec. Indian .,lead. Sci. BS1 181-185 Yashvir 1977 Induced polygenic variation in fruit length of Abelmoschus esculentus Moench. with

different mutagens; Geobios 4 179-182