Detection of Somaclonal Variation in Micropropagated and Acclimatized Plantlets of <i>Oryza sativa</i> MRQ 74 from Stem Explants

SALEH, A.; TAHA, R.M.; MAHMAD, N.; ELIAS, H.; HAMID, H.A.A.

doi:10.1590/S0100-83582019370100012

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Article • Planta daninha 37 • 2019 • https://doi.org/10.1590/S0100-83582019370100012 copy

Detection of Somaclonal Variation in Micropropagated and Acclimatized Plantlets of Oryza sativa MRQ 74 from Stem Explants

Detecção de Variação Somaclonal em Plantas Micropropagadas e Aclimatizadas de Oryza sativa MRQ 74 a Partir de Explantes do Caule

Authorship SCIMAGO INSTITUTIONS RANKINGS

ABSTRACT:

In plant tissue culture system, the excessive supply of plant growth hormones may inhibit further growth and development of the regenerants and sometimes it may induce somaclonal variation. In this study, the optimum concentration of plant growth hormones for in vitro regeneration of Oryza sativa L. cv. MRQ 74 was identified and subsequently the occurrence of somaclonal variation of regenerated plantlets was determined through cytological analysis of in vivo and in vitro grown plantlets. MS media supplemented with 0.1 mg L-1 BAP in combination with 0.1 mg L-1 NAA was found to be the optimum concentration for shoots and roots induction of Oryza sativa L. cv. MRQ 74. Plantlets derived from MS media supplemented with 0.5 mg L-1 2,4-D showed higher survival rate than that of MS media supplemented with 0.1 mg L-1 BAP in combination with 0.1 mg L-1 NAA. The plant height, number of leaf, leaf length and number of seeds per stalk of acclimatized plants were significantly lower (p<0.05) as compared with plants grown in vivo. Cytological analysis of in vivo and in vitro grown Oryza sativa L. cv. MRQ 74 revealed that the presence of 2,4-D in the culture media had significantly decreased the mitotic index (MI) and increased the ploidy level of the cell nuclei. The findings of the present study would be useful for plant breeders and biotechnologist since somaclonal variation provides a useful source of genetic variation for crop productivity and quality improvement.

Keywords:
plant growth hormones; Oryza sativa L. cv. MRQ 74; survival rate; agronomic parameters; mitotic index; ploidy level

Sociedade Brasileira da Ciência das Plantas Daninhas Departamento de Fitotecnia - DFT, Universidade Federal de Viçosa - UFV, 36570-000 - Viçosa-MG - Brasil, Tel./Fax::(+55 31) 3899-2611 - Viçosa - MG - Brazil
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[1] * Corresponding author: <azani783@salam.uitm.edu.my>

MS + Hormone (mg L^-1)		No. of shoots per explant (Mean ± SE)	No. of roots per explant (Mean ± SE)	Root length (cm)
BAP	NAA
0.0	0.0	2.30 ± 0.16 abc	5.27 ± 0.76 d	0.2 - 4.0
0.1	0.0	3.00 ± 0.27 cde	5.83 ± 1.18 d	0.3 - 4.0
	0.1	2.80 ± 0.23 bcde	25.33 ± 1.89 f	0.5 - 7.0
	0.5	1.97 ± 0.16 a	5.37 ± 0.74 d	0.2 - 4.5
	1.0	2.13 ± 0.21 ab	6.80 ± 0.70 d	0.4 - 4.5
0.5	0.0	3.13 ± 0.23 de	1.23 ± 0.25 a	0.4 - 2.7
	0.1	2.37 ± 0.15 abcd	5.53 ± 0.86 d	0.3 - 4.2
	0.5	3.03 ± 0.24 cde	2.57 ± 0.27 abc	0.2 - 0.7
	1.0	2.30 ± 0.18 abc	4.80 ± 0.58 cd	0.3 - 1.7
1.0	0.0	3.13 ± 0.26 de	1.27 ± 0.25 a	0.3 - 2.5
	0.1	2.70 ± 0.25 abcd	5.13 ± 0.66 d	0.3 - 4.3
	0.5	2.57 ± 0.22 abcd	9.80 ± 1.35 e	0.4 - 5.0
	1.0	3.53 ± 0.43 ef	4.40 ± 0.69 bcd	0.2 - 2.5
1.5	0.0	4.03 ± 0.31 f	1.37 ± 0.24 a	0.2 - 2.5
	0.1	2.30 ± 0.28 abc	2.67 ± 0.56 abc	0.3 - 2.0
	0.5	2.67 ± 0.24 abcd	2.13 ± 0.26 ab	0.2 - 2.3
	1.0	2.87 ± 0.18 bcde	1.87 ± 0.28 a	0.2 - 1.1

MS + Hormone (mg L^-1)		No. of shoots per explant (Mean ± SE)	No. of roots per explant (Mean ± SE)	Root length (cm)
NAA	kinetin
0.0	0.0	2.30 ± 0.16 bc	5.27 ± 0.76 a	0.2 - 4.0
0.0	0.5	2.57 ± 0.20 c	6.00 ± 0.58 ab	0.7 - 9.0
0.1	0.0	1.63 ± 0.21 a	24.37 ± 1.91 e	0.3 - 3.0
	0.1	2.40 ± 0.20 bc	8.63 ± 0.83 bc	0.2 - 3.2
	0.5	2.03 ± 0.18 abc	5.27 ± 0.69 a	0.2 - 2.0
0.5	0.0	2.10 ± 0.19 abc	9.90 ± 1.03 cd	0.3 - 5.5
	0.1	2.00 ± 0.19 abc	9.73 ±0.79 cd	0.4 - 5.0
	0.5	2.03 ± 0.19 abc	5.10 ± 0.62 a	0.2 - 2.5
1.0	0.0	1.80 ± 0.16 ab	12.17 ± 0.87 d	0.2 - 2.8
	0.1	2.07 ± 0.19 abc	10.30 ± 1.51 cd	0.3 - 5.7
	0.5	2.60 ± 0.18 c	8.77 ± 1.00 bc	0.4 - 3.0

MS + Hormone (mg L^-1)			No. of shoots per explant (Mean ± SE)	No. of roots per explant (Mean ± SE)	Root length (cm)
TDZ	NAA	BAP
0.0	0.0	0.0	2.30 ± 0.16 a	5.27 ± 0.76 b	0.2 - 4.0
0.1			8.23 ± 1.09 e	0.30 ± 0.16 a	0.2 - 0.7
0.2			6.47 ± 0.86 d	NR	NR
0.3			5.40 ± 0.73 cd	NR	NR
0.4			5.40 ± 0.60 cd	NR	NR
0.5			4.20 ± 0.33 abc	0.03 ± 0.03 a	0.2
0.1	0.1		3.17 ± 0.41 ab	0.03 ± 0.03 a	0.4
0.2	0.1		4.67 ±0.66 bcd	NR	NR
0.1	0.1	0.1	2.57 ± 0.22 a	0.07 ± 0.05 a	0.2 - 0.5

MS + Hormone (mg L^-1)		No. of shoots per explant (Mean ± SE)	No. of roots per explant (Mean ± SE)	Root length (cm)
2,4-D	Observation
0.0	No callus formation	2.30 ± 0.16 a	5.27 ± 0.76 a	0.2 - 4.0
0.5	Callus formation	4.51 ± 0.45 b	20.05 ± 2.34 b	0.3 - 5.0

Growing substrate	Survival rates of the plantlets (%)
Growing substrate	MS + 0.5 mg L^-1 2,4-D	MS + 0.1 mg L^-1 BAP + 0.1 mg L^-1 NAA
(4 weeks)
Black Soil	80.00 ± 2.52 b	3.33 ± 1.93 a
Red Soil	3.33 ± 0.33 a	0
Black + red soil	90.00 ± 1.53 c	10.00 ± 1.91 b
(8 weeks)
Black Soil	40.00 ± 1.15 a	0
Red Soil	0	0
Black + red soil	83.33 ± 1.20 b	0

Compound	Value (%)
Compound	Black soil	Red soil
Aluminium oxide (Al₂O₃)	9.158	43.165
Calcium oxide (CaO)	37.584	0.420
Cloride (Cl)	0.795	0.024
Iron(III) oxide	5.234	6.830
Lead(II) oxide (PbO)	0.120	0.010
Magnesium oxide (MgO)	1.598	0.000
Manganese(II) oxide (MnO)	0.151	0.021
Phosphorus pentoxide (P₂O₅)	1.758	0.108
Potassium oxide (K₂O)	2.380	0.447
Silicon dioxide (SiO₂)	35.445	l47.823
Sodium oxide (Na₂O)	0.286	0.053
Sulfur trioxide (SO₃)	5.162	0.183
Titanium dioxide (TiO₂)	1.094	0.773
Zinc oxide (ZnO)	0.080	0.015
Zirconium oxide (ZrO₂)	0.070	0.052

Parameter	In vivo	Acclimatized plantlet
Plant height (cm)	49.70 ± 1.08*	37.80 ± 0.91*
Number of leaves	8.07 ± 0.78*	5.37 ± 0.67*
Leaf length (cm)	40.83 ± 0.58*	26.80 ± 0.47*
Number of seeds per stalk	38.60 ± 2.07*	16.40 ± 0.81*

Treatment		Mitotic index, MI (%)	Number of chromosomes (mean)
In vivo		10.45 ± 0.31 b	24.00 ± 1.15 ab
In vitro	MS basal medium	10.22 ± 0.59 b	24.00 ± 2.65 ab
In vitro	MS medium supplemented with 0.5 mg L^-1 2,4-D	3.78 ± 0.22 a	22.00 ± 2.00 a

Treatment		Mean (µm²)
Treatment		Nucleus (N)	Cell (C)	Ratio (N/C)
In vivo		62.20 ± 1.96 b	191.00 ± 4.95 b	0.33 ± 0.03 b
In vitro	MS basal medium	48.92 ± 2.03 a	145.40 ± 6.37 a	0.34 ± 0.03 b
In vitro	MS + 0.5 mg L-1 2,4-D	64.67 ± 2.51 b	237.93 ± 19.71 b	0.29 ± 0.01 a

MS + hormones (mg L^-1)		Phase (%)
MS + hormones (mg L^-1)		G1	S	G2	> 4.8
In vivo		62.75 c	24.16 a	12.75 a	1.34 a
In vitro	MS basal medium	35.33 b	33.33 b	26.00 b	5.34 b
In vitro	MS + 0.5 mg L^-1 2,4-D	11.33 a	51.33 c	24.00 b	13.37 c