Micropropagation and assessment of somaclonal variation in <i>Galanthus transcaucasicus</i> in vitro plantlets

Asadi, Narges; Zarei, Hossein; Hashemi-Petroudi, Seyyed Hamidreza; Mousavizadeh, Seyyed Javad

doi:10.1590/2447-536X.v27i4.2320

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Scientific Article • Ornam. Hortic. 27 (4) • Oct-Dec 2021 • https://doi.org/10.1590/2447-536X.v27i4.2320 copy

Micropropagation and assessment of somaclonal variation in Galanthus transcaucasicus in vitro plantlets

Micropropagação e avaliação da variação somaclonal em plântulas de Galanthus transcaucasicus in vitro

Authorship SCIMAGO INSTITUTIONS RANKINGS

Abstract

In vitro culture of twin-scaling explants of Galanthus transcaucasicus with different concentrations of plant growth regulators (PGRs) including 0.5, 1, 2, 3, 4, 6, 8, and 10 mg L^-1 naphthaleneacetic acid (NAA) and 0.5, 1, 2, 3, and 4 mg L^-1 benzyladenine (BA) was studied. After 18 weeks, the number of regenerated bulblets and intensity of callus was measured. Subsequently, bulblets were transferred to a medium with 0.5, 1, 2, 3, and 4 mg L^-1 NAA and 0.5, 1, 2, 3, and 4 mg L^-1 BA and, after 15 weeks, the bulblets length and diameter were measured. The highest intensity of callus was obtained on 4 mg L^-1 NAA or 8 mg L^-1 NAA with 1 mg L^-1 BA. The highest number of regenerated bulblets was detected with 6 mg L^-1 NAA and 2 mg L^-1 BA. The highest diameter of bulblets occurred on four mgL^-1 NAA (9.4 mm), while the lowest was observed on 0.5 mg L^-1 BA (1.83 mm). The analysis of genetic variation using ISSR revealed that there was no somaclonal variation among the regenerated plants from BA and low level of NAA, but there was a significant somaclonal variation at high concentrations of NAA.

Keywords:
callus; embryogenesis; histology; ISSR; micropropagation; snowdrop

Growth regulator composition (mg L^-1)	Morphogenetic response	Number of regenerated bulblets/explant	Intensity of callus formation/explant (g)
2 NAA+1 BA	callus developing bulblets	2.40^de	0.30^ab
2 NAA+2 BA	direct bulblets	0.67^a	0.00^a
4 NAA+1 BA	callus developing bulblets	1.54^b	0.45^ab
4 NAA+2 BA	direct bulblets	1.55^b	0.00^a
6 NAA+1 BA	callus developing bulblets	1.64^bc	0.82^bc
6 NAA+2BA	callus developing bulblets	3.00^e	0.11^a
8 NAA+1 BA	callus developing bulblets	1.92^bcd	1.70^d
8NAA+2 BA	callus developing bulblets	1.33^ab	0.78^bc
10 NAA+1 BA	callus developing bulblets	2.31^cd	1.37^cd
10 NAA+2 BA	callus developing bulblets	1.33^ab	1.22^cd

NAA (mg L^-1)	Morphogenetic response	Number of regenerated bulblets/explant	Intensity of callus formation/explant (g)
0.5	callus developing bulblets	2.70^a	0.50^ab
1	callus developing bulblets	1.85^a	0.14^a
2	callus developing bulblets	1.66^a	1.17^bc
3	callus developing bulblets	1.66^a	1.75^c
4	callus developing bulblets	1.66^a	2.67^d

BA (mg L^-1)	Morphogenetic response	Number of regenerated bulblets/explant	Intensity of callus formation/explant (g)
0.5	direct bulblets	1.12^a	-
1	direct bulblets	1.16^a	-
2	direct bulblets	1.83^b	-
3	direct bulblets	2.00b	-
4	direct bulblets	2.87^c	-

Growth regulator composition (mg L^-1)	Average bulblet length (mm ± SD)	Median of bulblet length	Average bulblet diameter (mm ± SD)	Median of bulblet length diameter	Morphogenetic response
0 NAA, 0 BA	4.68 ± 0.39^a	4.7	1.84± 0.14^a	1.84	bulblets with well-visible shoot
0.5 NAA	5.18± 0.47^a	5.2	3.27± 0.59^b	3.27	bulblets with immature shoot
1 NAA	6.35 ± 0.64^bc	6.475	3.46 ± 0.73^b	3.48	bulblets with immature shoot
2 NAA	4.75 ± 0.59^a	4.58	4.44 ± 0.96^c	4.61	well-seen bulb scales and immature shoot
3 NAA	4.61 ± 0.41^a	4.61	5.74 ± 0.35^d	5.76	well-seen bulb scales and immature shoot
4 NAA	5.39 ± 0.33^ab	5.28	9.40 ± 0.58^e	9.295	well-seen bulb scales and immature shoot
0.5 BA	5.6 ± 0.31^abc	5.62	1.83 ± 0.87^a	1.535	immature bulblets with well-visible shoot
1 BA	6.59 ± 0.73^cd	6.48	2.53 ± 0.48^ab	2.35	mature bulblets with well-visible shoot
2 BA	6.54 ± 0.44^cd	6.52	3.17 ± 0.57^b	3.155	mature bulblets with well-visible shoot
3 BA	7.44 ± 0.71^d	7.7	3.46 ± 0.45^b	3.40	mature bulblets with well-visible shoot
4 BA	11.24 ± 0.66^e	11.16	3.47 ± 0.34^b	3.44	mature bulblets with well-visible shoot

Sl. No.	Primer Code	Sequence (5’-3’)	Tm (ºC)	Total bands	No. of polymorphic bands	Polymorphism (%)	Range (bp)
1	ISSR-2	GAGAGAGAGAGAGAGAGAC	57	8	8	100.00	700-2500
2	ISSR-7	GAGAGAGAGAGAGAGAC	52	8	7	87.50	550-1750
3	ISSR-10	AGAGAGAGAGAGAGAGC	52	10	10	100.00	500-1500
4	ISSR-14	TCTCTCTCTCTCTCTCG	52	5	4	80.00	700-2000
5	ISSR-15	ACACACACACACACACG	52	13	10	79.92	300-2000
6	ISSR-17	ACACACACACACACACC	52	6	6	100.00	250-1750
7	ISSR-19	ATCATCATCATCATCATCC	52	6	4	80.00	600-1500
Total	-	-	-	56	49	-	-
Mean				8	7	89.63

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[1] * Corresponding author: n.asadi82@yahoo.com