Micropropagation of Mountain Mulberry (Morus bombycis Koidz.) ‘Kenmochi’ on Cytokinin-Free Medium
Abstract
:1. Introduction
2. Materials and Methods
2.1. Initiation of In Vitro Cultures
2.2. Multiplication Stage (Main Experiments)
2.3. Ex Vitro Rooting of Shoots (Additional Experiment)
2.4. Statistical Analyses
3. Results
3.1. Initiation of In Vitro Cultures
3.2. Multiplication Stage (Main Experiments)
3.3. Ex Vitro Rooting of Shoots (Additional Experiment)
4. Discussion
5. Conclusions
Author Contributions
Funding
Conflicts of Interest
Ethical Statements
Abbreviations
BA | 6-Benzylaminopurine |
IBA | Indole-3-butyric acid |
MS | Murashige and Skoog medium [14] |
WPM | Woody Plant Medium [15] |
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Media Constituents | “AxB” Medium (Standard) | “SNC” Medium |
---|---|---|
Macronutrients | 50% MS 1 | |
N salts | 100% MS | |
K salts | 100% MS | |
P salts | 100% MS | |
Ca salts | 125% MS | |
Mg salts | 125% MS | |
Micronutrients | 100% MS | 50% MS |
Vitamins | 100% WPM2 | 100% WPM |
Myo-inositol | 100 mg L−1 | 100 mg L−1 |
Sucrose | 30 g L−1 | 15 g L−1 |
BA | 1.5 mg L-1 | - |
IBA | - | 0.05 mg L−1 |
Agar | 7 g L−1 | 7 g L−1 |
pH | 5.8 | 5.8 |
Source of Explants | No of Survived Explants (S) [%] | No of Explants which Started to Grow (G) [%] | Efficiency of Initiation (S x G) [%] | |||
---|---|---|---|---|---|---|
“SNC” | “AxB” | “SNC” | “AxB” | “SNC” | “AxB” | |
field tree | 52 a,A | 100 b,B | 38 a,A | 100 b,B | 21 a,A | 100 b,B |
nursery plants | 100 b,B | 92 a,A | 64 a,B | 82 a,A | 64 a,B | 75 a,A |
Medium/ No of Subculture | Explant Type | No of Growing Cultures [%] | Number of Shoots [pcs] | Length of the Longest Shoot [cm] | Total Length of Shoots [cm] | Callus Size [cm] |
---|---|---|---|---|---|---|
SNC/1 | ST | 91.7 a | 1.0 a | 5.9 b | 6.0 a | 0.0 a |
AxB/1 | ST | 91.7 a | 1.8 b* | 3.2 a | 4.9 a | 2.3 b |
SNC/2 | 1N | 96.9 a | 1.0 a | 7.4 b | 7.5 b | 0.0 a |
AxB/2 | xN | 100 a | 1.6 b | 2.0 a | 2.6 a | 1.6 b |
SNC/3 | 1N | 100 a | 1.0 a | 5.2 a | 5.2 a | 0.0 a |
AxB/3 | xN | 100 a | 1.7 b | 5.0 a | 7.0 b | 1.1 b |
SNC/4 | 1N | 86.3 a | 1.0 a | 5.9 b | 6.1 b | 0.0 a |
AxB/4 | xN | 100 a | 1.6 b | 1.9 a | 2.6 a | 1.5 b |
SNC/5 | 1N | 100 a | 1.0 a | 5.5 b | 5.5 a | 0.0 a |
AxB/5 | xN | 95.6 a | 2.1 b | 3.2 a | 5.3 a | 1.7 b |
SNC/6 | 1N | 97.5 a | 1.0 a | 2.8 b | 2.8 a | 0.0 a |
AxB/6 | xN | 91.5 a | 1.4 b | 1.9 a | 2.5 a | 0.9 b |
SNC (mean) | 95.4 - | 1.0 - | 5.5 - | 5.5 - | 0.0 - | |
AxB (mean) | 96.5 - | 1.7 - | 2.9 - | 4.2 - | 1.5 - |
Type of Culture | No of Growing Cultures [%] | Number of Shoots [pcs] | Length of the Longest Shoot [cm] | Total Length of Shoots [cm] | Callus Size [cm] |
---|---|---|---|---|---|
Old cultures after one harvest of shoots | 86.3 a | 1.3 a | 2.2 a | 2.7 a | 0.0 a |
New culture | 100 a | 1.0 a | 5.5 b | 5.5 b | 0.0 a |
Origin of Explant/ No of Subculture | Explant Type | No of Growing Cultures [%] | Number of Shoots [pcs] | Length of the Longest Shoot [cm] | Total Length of Shoots [cm] | Callus Size [cm] |
---|---|---|---|---|---|---|
from SNC/2 * | 1N ** | 93.8 a | 1.3 a | 2.7 b | 3.0 a | 2.0 b |
from AxB/2 | xN | 100 a | 1.6 b | 2.0 a | 2.6 a | 1.6 a |
from SNC/3 | 1N | 100 a | 1.2 a | 3.3 a | 3.7 a | 1.4 b |
from AxB/3 | xN | 100 a | 1.7 b | 5.0 a | 7.0 b | 1.1 a |
from SNC/5 | 1N | 100 a | 2.1 a | 1.7 a | 2.8 a | 1.9 b |
from AxB/5 | xN | 95.6 a | 2.1 a | 3.2 b | 5.3 b | 1.7 a |
from SNC/6 | 1N | 100 a | 1.6 a | 2.1 a | 2.9 a | 0.9 a |
from AxB/6 | xN | 91.5 a | 1.4 a | 1.9 a | 2.5 a | 0.9 a |
from SNC (mean) | 98.5 - | 1.6 - | 2.5 - | 3.1 - | 1.6 - | |
from AxB (mean) | 96.8 - | 1.7 - | 3.0 - | 4.4 - | 1.3 - |
Explant Type | No of Growing Cultures [%] | Number of Shoots [pcs] | Length of the Longest Shoot [cm] | No of Rooted Shoots [%] | Callus Size [cm] |
---|---|---|---|---|---|
1N | 96.9 a | 1.0 a | 2.9 a | 100 a | 0.0 a |
2N | 100 a | 1.0 a | 2.5 a | 100 a | 0.0 a |
Explant Type | No of Growing Cultures [%] | Number of Shoots [pcs] | Length of the Longest Shoot [cm] | Total Length of Shoots [cm] | Callus Size [cm] |
---|---|---|---|---|---|
xN | 95.8 a | 1.4 a | 1.7 a | 2.1 a | 0.9 a |
ST | 91.3 a | 2.2 b | 3.4 b | 5.4 b | 0.8 a |
Origin of Microshoots | No of Rooted Shoots [%] | No of Shoots [pcs] | Length of the Longest Shoot [cm] | Total Length of Shoots [cm] | Length of Leaf Blade [cm] | Width of Leaf Blade [cm] | Relative Chlorophyll Content [SPAD] |
---|---|---|---|---|---|---|---|
SNC | 100 a | 1.1 a | 6.0 a | 6.2 a | 3.5 a | 2.6 a | 23.3 a |
AxB | 100 a | 1.3 a | 6.1 a | 6.8 a | 3.3 a | 2.7 a | 24.2 a |
Method of Micropropagation | Axillary Branching (“AxB”) | Single-Node Culture (“SNC”) |
---|---|---|
Cost of media | higher | lower (reduced dose of sucrose and mineral nutrients) |
Efficiency of micropropagation | ||
Initiation of in vitro cultures (explants from young, nursery plants) | easy | easy |
Initiation of in vitro cultures (explants from adult, field trees) | possible | possible although hard to get |
Number of shoots | significantly higher | significantly lower |
Length of shoots | significantly lower | significantly higher |
Number of nodes on shoot | similar | similar |
Estimated multiplication rate * | lower: 4 (2 × “ST”+ 2 × “xN” explants) | higher: 5 (1 × “ST” + 4 × “1N” explants) |
Rooting and acclimation in vivo | similar | similar |
Quality of cultures during multiplication | ||
Growth of callus | too intensive | not observed |
Physiological disorders (shoot-tip necrosis, vitrification, chlorosis, etc.) | sometimes observed | not observed |
Development of adventitious shoots | rather not observed (overgrown callus complicates identification) | not observed |
Risk of somaclonal variation | low | very low |
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Litwińczuk, W.; Jacek, B. Micropropagation of Mountain Mulberry (Morus bombycis Koidz.) ‘Kenmochi’ on Cytokinin-Free Medium. Plants 2020, 9, 1533. https://doi.org/10.3390/plants9111533
Litwińczuk W, Jacek B. Micropropagation of Mountain Mulberry (Morus bombycis Koidz.) ‘Kenmochi’ on Cytokinin-Free Medium. Plants. 2020; 9(11):1533. https://doi.org/10.3390/plants9111533
Chicago/Turabian StyleLitwińczuk, Wojciech, and Beata Jacek. 2020. "Micropropagation of Mountain Mulberry (Morus bombycis Koidz.) ‘Kenmochi’ on Cytokinin-Free Medium" Plants 9, no. 11: 1533. https://doi.org/10.3390/plants9111533