Boron on <i>in vitro</i> growth and enzymatic activity of Blueberry

Boron (B) is essential for plants metabolism and most culture mediums use the same concentration, but in different quantities this nutrient may provoke growth alterations. Thus, the purpose of this study was to examine the effect of B on in vitro growth of blueberry in three experiments. The first experiment of multiplication (evaluated at 90 days) used 3 concentrations of 2-isopentenyladenine-2iP with 4 concentrations of boric acid-BA (factorial scheme 3x4). The second used 4 concentrations of BA and 1 concentration of 2iP and the third (rooting) used 4 concentrations of BA and 1 concentration of indolbutyric acid-IBA both evaluated at 180 days (unifactorial scheme). All the experiments had 4 replicates with explants. 6.2 mg L-1 of B and 5.0 mg L-1 of 2iP generated the highest shoot quantity (18.4, 25.5 respectively). From the interaction of these concentrations, there was the highest activity of POD and PPO enzymes. Under B deficit was seen a larger number of shoot-tip necrosis (9), red leaves (31) and high activity of the PAL, IAAO and POD enzymes. With IBA the correlation between IAAO and the roots’ growth was positive, showing that blueberry rooting depends of both B and IAAO regulation.

Index terms
Vaccinium ashei Reade; Boric acid; Plant propagation

Authorship

Jessé Neves dos Santos

MSc. in Agronomy by the State University of Ponta Grossa (UEPG) Ponta Grossa – PR, Brazil. Email: jessenevessantos@hotmail.com State University of Ponta GrossaBrasilPonta Grossa, PR, Brasil MSc. in Agronomy by the State University of Ponta Grossa (UEPG) Ponta Grossa – PR, Brazil. Email: jessenevessantos@hotmail.com

http://orcid.org/0000-0002-4228-8298

Ricardo Antonio Ayub

Agronomist, PhD, Professor Researcher, State University of Ponta Grossa (UEPG), Ponta Grossa – PR, Brazil. Email: rayub@uepg.br State University of Ponta GrossaBrasilPonta Grossa, PR, Brasil Agronomist, PhD, Professor Researcher, State University of Ponta Grossa (UEPG), Ponta Grossa – PR, Brazil. Email: rayub@uepg.br

http://orcid.org/0000-0003-3240-8417

Isabela Letícia Pessenti

http://orcid.org/0000-0002-5176-3134

André Belmont Pereira

Agronomist, PhD, Professor Researcher, State University of Ponta Grossa (UEPG), Ponta Grossa – PR, Brazil. Email: abelmont@uepg.br State University of Ponta GrossaBrasilPonta Grossa, PR, Brasil Agronomist, PhD, Professor Researcher, State University of Ponta Grossa (UEPG), Ponta Grossa – PR, Brazil. Email: abelmont@uepg.br

http://orcid.org/0000-0002-1673-9841

SCIMAGO INSTITUTIONS RANKINGS

Figures | Tables

Figures (7)
Tables (3)

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Figure 1
Influence of boric acid and 2iP on the development of aerial part of blueberry at 90-day cultivation. A and B – callus diameter, C and D – number of shoots, E and F – number of leaves, G – length of shoots, and H – fresh weight. Averages followed by the same letters among concentrations of 2iP did not statistically differ by means of the Tukey test at 5% reliability; *significant at 5% reliability; **significant at 1% reliability; ns – non-significant; A and B had data transformed by the formula prior to statistical analysis; C, D, E and F had data transformed by the formula log (x+1) prior to statistical analysis.

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Figure 2
Plants of blueberry. A, B and C – Plants grown at 90-day cultivation in a medium containing 0, 2.5 and 5.0 mg L-1 of 2iP, respectively, without boric acid. D and E – symptoms evidencing deficiency of boron at 180-day in vitro cultivation. F, G, H and I – plants grown at 180-day cultivation in a medium containing 0, 3.1, 6.2, and 9.3 mg L^-1 of boric acid, respectively, with 5.0 mg L^-1 of 2iP concentration. J, K, L and M – plants grown for 180 days in a medium containing 0, 3.1, 6.2, and 9.3 mg L^-1 of boric acid, respectively, with 2.5 mg L^-1 of IBA concentration.

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Figure 3
Influence of boric acid and 2iP on the leaf (L) enzymatic activity at 90-day in vitro cultivation. A – polyphenol oxidase; B – peroxidase; C – phenylalanine ammonia lyase. Averages followed by the same capital letters among BA concentrations along with small letters among 2iP concentrations did not statistically differ by means of the Tukey test at 5% reliability.

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Figure 4
Influence of boric acid on the aerial part of blueberry grown for 180 days in a medium containing 5.0 mg L^-1 of 2iP. A – Length of shoots; B – number of red leaves; C – number of shoot-tips necrosis; D – amount of boron in aerial part; *significant at 5% reliability; **significant at 1% reliability.

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Figure 5
Influence of boric acid on the aerial part of blueberry grown for 180 days in a medium containing 5.0 mg L^-1 of 2iP. A – Length of shoots; B – number of red leaves; C – number of shoot-tips necrosis; D – amount of boron in aerial part; *significant at 5% reliability; **significant at 1% reliability.

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Figure 6
Influence of boric acid on the growth of blueberry grown for 180 days in a medium containing 2.5 mg L^-1 of IBA. A – length of shoots; B – callus diameter; C – number of leaves; D – fresh weight; E – rooting percentage. **significant at 1% reliability.

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Figure 7
Influence of boric acid on the enzymatic activity of blueberry grown in a medium containing 2.5 mg L^-1 of IBA. A and B – activity of PPO and PAL enzymes in the callus, respectively; C, D and E – activity of PAL, POD and IAAO enzymes in the leaf, respectively; ** significant at 1% reliability.

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Table 1
Pearson correlation study taking into account the following variables from experiment I related to in vitro multiplication: BA – boric acid; C – cytokinin; PPOL – leaf PPO; PALL – leaf PAL; PODL – leaf POD; IAAOL – leaf IAAO; LS – length of shoots; FW – fresh weight; CD – callus diameter; NL – number of leaves; NS – number of shoots.

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Table 2
Pearson correlation study taking into account the following variables from experiment II related to in vitro multiplication: BA – boric acid; LS – length of shoots; CD – callus diameter; NL – number of leaves; NRL – number of red leaves; NNBT – number of necrosed bud-tips; NS – number of shoots; FW – fresh weight; NR12 – number of roots of first and second order; NR3 – number of roots of third order; LLR – Length of the longest root; ABAP – amount of boron in aerial part; PPOL – leaf PPO; PALL – leaf PAL; PODL – leaf POD; IAAOL – leaf IAAO; PPOC – callus PPO; PALC – callus PAL; PODC – callus POD; IAAOC – callus IAAO.

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Table 3
Pearson correlation study taking into account the following variables from experiment III related to in vitro multiplication: BA – boric acid; LS – length of shoots; CD – callus diameter; NL – number of leaves; NS – number of shoots; FM – fresh mass; NR – number of roots; LLR – length of the longest root; RP – rooting percentage; PPOL – leaf PPO; PALL – leaf PAL; PODL – leaf POD; IAAOL – leaf IAAO; PPOC – callus PPO; PALC – callus PAL; PODC – callus POD; IAAOC – callus IAAO.

Figure 1 Influence of boric acid and 2iP on the development of aerial part of blueberry at 90-day cultivation. A and B – callus diameter, C and D – number of shoots, E and F – number of leaves, G – length of shoots, and H – fresh weight. Averages followed by the same letters among concentrations of 2iP did not statistically differ by means of the Tukey test at 5% reliability; *significant at 5% reliability; **significant at 1% reliability; ns – non-significant; A and B had data transformed by the formula prior to statistical analysis; C, D, E and F had data transformed by the formula log (x+1) prior to statistical analysis.

Figure 2 Plants of blueberry. A, B and C – Plants grown at 90-day cultivation in a medium containing 0, 2.5 and 5.0 mg L-1 of 2iP, respectively, without boric acid. D and E – symptoms evidencing deficiency of boron at 180-day in vitro cultivation. F, G, H and I – plants grown at 180-day cultivation in a medium containing 0, 3.1, 6.2, and 9.3 mg L^-1 of boric acid, respectively, with 5.0 mg L^-1 of 2iP concentration. J, K, L and M – plants grown for 180 days in a medium containing 0, 3.1, 6.2, and 9.3 mg L^-1 of boric acid, respectively, with 2.5 mg L^-1 of IBA concentration.

Figure 3 Influence of boric acid and 2iP on the leaf (L) enzymatic activity at 90-day in vitro cultivation. A – polyphenol oxidase; B – peroxidase; C – phenylalanine ammonia lyase. Averages followed by the same capital letters among BA concentrations along with small letters among 2iP concentrations did not statistically differ by means of the Tukey test at 5% reliability.

Figure 4 Influence of boric acid on the aerial part of blueberry grown for 180 days in a medium containing 5.0 mg L^-1 of 2iP. A – Length of shoots; B – number of red leaves; C – number of shoot-tips necrosis; D – amount of boron in aerial part; *significant at 5% reliability; **significant at 1% reliability.

Figure 5 Influence of boric acid on the aerial part of blueberry grown for 180 days in a medium containing 5.0 mg L^-1 of 2iP. A – Length of shoots; B – number of red leaves; C – number of shoot-tips necrosis; D – amount of boron in aerial part; *significant at 5% reliability; **significant at 1% reliability.

Figure 6 Influence of boric acid on the growth of blueberry grown for 180 days in a medium containing 2.5 mg L^-1 of IBA. A – length of shoots; B – callus diameter; C – number of leaves; D – fresh weight; E – rooting percentage. **significant at 1% reliability.

Figure 7 Influence of boric acid on the enzymatic activity of blueberry grown in a medium containing 2.5 mg L^-1 of IBA. A and B – activity of PPO and PAL enzymes in the callus, respectively; C, D and E – activity of PAL, POD and IAAO enzymes in the leaf, respectively; ** significant at 1% reliability.

Table 1 Pearson correlation study taking into account the following variables from experiment I related to in vitro multiplication: BA – boric acid; C – cytokinin; PPOL – leaf PPO; PALL – leaf PAL; PODL – leaf POD; IAAOL – leaf IAAO; LS – length of shoots; FW – fresh weight; CD – callus diameter; NL – number of leaves; NS – number of shoots.

	BA	C	PPOL	PALL	PODL	IAAOL	LS	FW	CD	NL	NS
BA	1.00	ns	0.60*	ns	ns	ns	ns	ns	ns	ns	ns
C	-	1.00	0.65**	0.90**	0.59*	0.64**	ns	0.95**	0.97**	0.84**	0.87**
PPOL	-	-	1.00	0.72**	0.71**	0.56*	ns	0.58*	0.54*	0.62*	0.80**
PALL	-	-	-	1.00	0.54*	0.56*	ns	0.76**	0.82**	0.87**	0.76**
PODL	-	-	-	-	1.00	0.66**	ns	0.57*	ns	0.60*	0.72**
IAAOL	-	-	-	-	-	1.00	ns	0.61*	0.57*	ns	0.72**
LS	-	-	-	-	-	-	1.00	ns	ns	ns	ns
FW	-	-	-	-	-	-	-	1.00	0.94*	0.67**	0.88**
CD	-	-	-	-	-	-	-	-	1.00	0.77**	0.85**
NL	-	-	-	-	-	-	-	-	-	1.00	0.68**
NS	-	-	-	-	-	-	-	-	-	-	1.00

^*
significant at 5% reliability; **significant at 1% reliability; ns – non-significant.

Table 2 Pearson correlation study taking into account the following variables from experiment II related to in vitro multiplication: BA – boric acid; LS – length of shoots; CD – callus diameter; NL – number of leaves; NRL – number of red leaves; NNBT – number of necrosed bud-tips; NS – number of shoots; FW – fresh weight; NR12 – number of roots of first and second order; NR3 – number of roots of third order; LLR – Length of the longest root; ABAP – amount of boron in aerial part; PPOL – leaf PPO; PALL – leaf PAL; PODL – leaf POD; IAAOL – leaf IAAO; PPOC – callus PPO; PALC – callus PAL; PODC – callus POD; IAAOC – callus IAAO.

	BA	LS	CD	NL	NRL	NNBT	NS	FW	NR12	NR3	LLR	ABAP	PPOL	PALL	PODL	IAAOL	PPOC	PALC	PODC	IAAOC
BA	1	ns	ns	ns	-0.85**	-0.66**	ns	ns	0.74**	0.72**	0.78**	0.98**	ns	ns	ns	-0.69**	-0.72**	ns	-0.82**	ns
LS	-	1	ns	ns	-0.65**	-0.51*	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns	-0.59*	ns	-0.55*	ns
CD	-	-	1	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns
NL	-	-	-	1	ns	ns	0.76**	0.55*	0.53*	0.52*	0.53*	ns	-0.69**	ns	-0.52*	ns	ns	ns	-0.52*	ns
NRL	-	-	-	-	1	0.80**	ns	ns	-0.94**	-0.90**	-0.92**	-0.83**	ns	0.67**	0.89**	0.74**	0.96**	ns	0.95**	ns
NNBT	-	-	-	-	-	1	ns	ns	-0.77**	-0.78**	-0.81	-0.62	ns	0.50*	0.78**	0.70**	0.85**	ns	0.83**	ns
NS	-	-	-	-	-	-	1	0.67**	ns	0.51*	ns	ns	-0.66**	ns	ns	ns	ns	ns	ns	ns
FW	-	-	-	-	-	-	-	1	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns	ns
NR12	-	-	-	-	-	-	-	-	1	0.97**	0.93**	0.71**	-0.62*	-0.61*	-0.87**	-0.65**	-0.93**	ns	-0.96**	ns
NR3	-	-	-	-	-	-	-	-	-	1	0.93**	0.70**	-0.70**	-0.54*	-0.89**	-0.68**	-0.90**	ns	-0.96**	ns
LLR	-	-	-	-	-	-	-	-	-	-	1	0.78**	-0.64**	-0.64**	-0.91**	-0.72**	-0.93**	ns	-0.93**	ns
ABAP	-	-	-	-	-	-	-	-	-	-	-	1	ns	ns	-0.69**	-0.88**	-0.71**	ns	-0.77**	ns
PPOL	-	-	-	-	-	-	-	-	-	-	-	-	1	ns	0.53*	ns	ns	-0,62*	-0.58*	ns
PALL	-	-	-	-	-	-	-	-	-	-	-	-	-	1	0.68**	ns	0.64**	ns	0.65**	ns
PODL	-	-	-	-	-	-	-	-	-	-	-	-	-	-	1	0.64**	0.92**	ns	0.90**	ns
IAAOL	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	1	0.67**	ns	0.78**	ns
PPOC	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	1	ns	0.92**	ns
PALC	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	1	ns	-0.74**
PODC	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	1	ns
IAAOC	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	1

^*
significant at 5% reliability;
^**
significant at 1% reliability; ns – non-significant.

Table 3 Pearson correlation study taking into account the following variables from experiment III related to in vitro multiplication: BA – boric acid; LS – length of shoots; CD – callus diameter; NL – number of leaves; NS – number of shoots; FM – fresh mass; NR – number of roots; LLR – length of the longest root; RP – rooting percentage; PPOL – leaf PPO; PALL – leaf PAL; PODL – leaf POD; IAAOL – leaf IAAO; PPOC – callus PPO; PALC – callus PAL; PODC – callus POD; IAAOC – callus IAAO.

	BA	LS	CD	NL	NS	FW	NR	LLR	RP	PPOL	PALL	PODL	IAAOL	PPOC	PALC	PODC	IAAOC
BA	1	ns	-0.74**	ns	ns	ns	ns	ns	0.78	ns	-0.60*	-0.71**	ns	ns	ns	ns	ns
LS	-	1	ns	0.62*	0.74**	ns	ns	Ns	ns	-0.68**	ns	-0.56*	ns	0.66	ns	ns	ns
CD	-	-	1	ns	ns	ns	ns	Ns	-0.59*	ns	ns	ns	ns	ns	0.54*	ns	ns
NL	-	-	-	1	ns	-0.67**	ns	Ns	ns	ns	ns	-0.78**	0.65**	0.74**	0.72**	ns	ns
NS	-	-	-	-	1	ns	ns	Ns	ns	-0.51**	ns	ns	ns	ns	ns	ns	ns
FW	-	-	-	-	-	1	ns	Ns	ns	ns	ns	ns	-0.62**	-0.74	-0.64**	ns	ns
NR	-	-	-	-	-	-	1	0.60*	ns	ns	ns	ns	ns	ns	ns	ns	ns
LLR	-	-	-	-	-	-	-	1	ns	-0.59	ns	ns	ns	ns	ns	ns	ns
RP	-	-	-	-	-	-	-	-	1	ns	ns	-0.53*	0.50*	ns	ns	ns	ns
PPOL	-	-	-	-	-	-	-	-		1	ns	ns	ns	ns	ns	ns	ns
PALL	-	-	-	-	-	-	-	-	-	-	1	0.54*	ns	ns	ns	ns	ns
PODL	-	-	-	-	-	-	-	-	-	-	-	1	-0.73**	-0.51*	ns	ns	ns
IAAOL	-	-	-	-	-	-	-	-	-	-	-	-	1	0.72**	ns	ns	ns
PPOC	-	-	-	-	-	-	-	-	-	-	-	-	-	1	0.67**	ns	ns
PALC	-	-	-	-	-	-	-	-	-	-	-	-	-	-	1	ns	ns
PODC	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	1	ns
IAAOC	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	1

^*
significant at 5% reliability;
^**
significant at 1% reliability; ns – non-significant

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Boron on <i>in vitro</i> growth and enzymatic activity of Blueberry

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[TFN0101] ^*
significant at 5% reliability; **significant at 1% reliability; ns – non-significant.