quantification-endogenous-auxin-cytokinin-during-internode-culture

Quantification of Endogenous Auxin and Cytokinin During Internode Culture of Ipecac

Adventitious shoot formation is an important technique for the propagation of economically important crops and for the regeneration of transgenic plants. ...

Toyo University

Toyo university

Adventitious root cultures of Physochlaina physaloides were established and their growth and tropane alkaloid production were investigated. Although the roots cultured in Murashige-skoog liquid medium containing 1.0-3.0 mg/l IBA grew well, the addition of lower concentrations of auxins was more suitable for alkaloid production. The best alkaloid yield was obtained with 0.01 mg/l NAA in 1/2 MS liquid medium. In addition, the alkaloid differences between the cultured roots and the regenerated plant cultivated in the soil was studied. Cultured roots in this condition showed higher level of alkaloids when compared to leaf, stem and roots from the regenerated plant (3 months in field conditions).

Tropane alkaloids in adventitious root cultures of Physochlaina physaloides.

Adventitious and hairy root cultures of Physochlaina physaloides were established. These roots grew well and produced high amounts of tropane alkaloids (particularly hyoscyamine and 6 beta-hydroxyhyoscyamine) in auxin-free culture medium. The effects of basal media and temperature on the growth and alkaloid production of these roots were investigated. Both root cultures produced highest amount of tropane alkaloids in B5 medium though the optimum temperature for hairy roots were lower than that for adventitious roots.

Tropane alkaloids in auxin-independent root cultures of Physochlaina physaloides.

The effect of fertilizer on the yield and yield components of opium from Papaver somniferum L.cv.Ikkanshu were investigated from 1993 to 1995. The weight of opium per one capsule was tendency to increase in the order of peat moss > pulverized charcoal > compound fertilizer, but the differences among the results were not significant. Furthermore, no difference in the capsule weight per 100 m2 and the yields of opium and alkaloids per 100 m2 were detected. The significant difference was only observed when the data were compared among the weights and yields of different years. The difference among the yield of opium was mainly due to the weight of opium per one capsule. The weight of opium per one capsule showed a high correlation with capsule husk weight. The alkaloids contents in opium obtained with a different fertilizer application showed no difference. Morphine content at the first lancing was about 11%, and the value decreased with the order of lancing. On the contrary, the codeine and the thebaine content did not change during lancing and the value were 6-7% and 3%, respectively. The papaverine and the noscapine content decreased in the order of lancing.

[Influence of fertilizers on yield and yield components of opium from Papaver somniferum Linn].

Lithospermum erythrorhizon shoots, cultured on phytohormone-free Murashige and Skoog solid medium, produced shikonin derivatives, whereas shoots cultured in well-ventilated petri dishes, produced small amount. Analysis by gas chromatography revealed the presence of ethylene in non-ventilated petri dishes where the shoots, producing shikonin derivatives, were cultured. Therefore, the possible involvement of ethylene in shikonin biosynthesis of shoot cultures was investigated. Treatment of ethylene or the ethylene precursor, 1-aminocyclopropane-1-carboxylic acid, resulted in increasing shikonin derivatives contents in cultured shoots. Silver ion, an ethylene-response inhibitor, or aminoethoxyvinylglycine, an ethylene biosynthesis inhibitor, decreased production of shikonin derivatives in cultured shoots. Our results indicate that ethylene is one of the regulatory elements of shikonin biosynthesis in L. erythrorhizon shoot culture.

Ethylene induced shikonin biosynthesis in shoot culture of Lithospermum erythrorhizon.

Atropa belladonna hairy roots (clone M8) were successfully cryopreserved by using the vitrification method. A. belladonna hairy root tips were precultured on a half strength of Murashige and Skoog (MS) solid medium with 0.1 mg per L 2,4-D or without phytohormone for 1 day, and then dehydrated with PVS2 solution for 15 minutes prior to immersion into liquid nitrogen for 1 day, 1 week, 1 month and 3 months. Hairy root tips kept in liquid nitrogen were rapidly thawed at 36 degree C in a water bath. The root tips were recultured on half strength MS medium. The hairy root tips, precultured with 2,4-D before cryopreservation, showed a higher survival rate than those precultured without phytohormone. The hairy root tips, precultured with 2,4-D, showed an average survival rate of 83 percent. There was no significant difference in the viability of the hairy roots cryopreserved for different periods. The regrowth of cryopreserved hairy roots was similar to that of untreated hairy roots and tropane alkaloid productivity became stable after 4th subculture. PCR analysis of hairy roots demonstrated the conservation of the T-DNA in cryopreserved hairy roots. These results indicate that cryopreservation by vitrification method is useful to preserve A.belladonna hairy root clone M8.

Characteristics of Atropa belladonna hairy roots cryopreserved by vitrification method.

Gynura bicolor DC. is a traditional vegetable in Japan. G. bicolor grown in the field has adaxial sides of leaves that are green and abaxial sides that are reddish purple. It has been reported that the responsible reddish purple pigments are anthocyanins, which are acylated and highly stable. We have reported that cultured G. bicolor plantlets treated with methyl jasmonate (MJ) exhibited anthocyanin accumulation in roots, and this was affected by light irradiation. In the present study, to clarify this accumulation induced by MJ treatment, we isolated anthocyanin biosynthesis and regulatory genes from G. bicolor. Expression analysis revealed up-regulated expression of flavonoid biosynthesis genes, GbCHS, GbCHI, GbDFR and GbANS. Furthermore, it was shown that isolated regulatory genes, GbMYB1 and GbMYC1, were also up-regulated by MJ treatment. In addition, it was shown that co-expression of GbMYB1 and GbMYC1 could activate GbDFR and GbANS gene promoters in transient assays with tobacco protoplasts. These results strongly indicate that GbMYB1 and GbMYC1 coordinately regulate flavonoid biosynthetic genes induced by MJ treatment, and thereby cause anthocyanin accumulation in roots.

Co-expression of GbMYB1 and GbMYC1 induces anthocyanin accumulation in roots of cultured Gynura bicolor DC. plantlet on methyl jasmonate treatment.

Nakajimana (Brassica rapa L. cv. nakajimana), of the family Brassicaceae, is a traditional vegetable in Japan. Three isothiocyanates and five cyanides in the leaves of nakajimana were identified by gas chromatography (GC) and GC-mass spectrometry (GC-MS), and their kinetic changes using heat-treatment (temperature and time) were investigated. In addition, myrosinase activity of extracts prepared from fresh nakajimana leaf was determined. In crushed heat-treated leaves of nakajimana (70 °C for 30 s), formation of isothiocyanates and myrosinase activity increased, whereas formation of 1-cyano-3,4-epithiobutane and 1-cyano-4,5-epithiopentane decreased. Heat-treatment can significantly alter the content of potentially beneficial compounds in nakajimana, and ingestion of suitable isothiocyanates for human health may be better facilitated by mild boiling.

Kinetic changes in glucosinolate-derived volatiles by heat-treatment and myrosinase activity in nakajimana (Brassica rapa L. cv. nakajimana).

Strigolactones (SLs) act as plant hormones that inhibit shoot branching and stimulate secondary growth of the stem, primary root growth, and root hair elongation. In the moss Physcomitrella patens, SLs regulate branching of chloronemata and colony extension. In addition, SL-deficient and SL-insensitive mutants show delayed leaf senescence. To explore the effects of SLs on leaf senescence in rice (Oryza sativa L.), we treated leaf segments of rice dwarf mutants with a synthetic SL analogue, GR24, and evaluated their chlorophyll contents, ion leakage, and expression levels of senescence-associated genes. Exogenously applied GR24 restored normal leaf senescence in SL-deficient mutants, but not in SL-insensitive mutants. Most plants highly produce endogenous SLs in response to phosphate deficiency. Thus, we evaluated effects of GR24 under phosphate deficiency. Chlorophyll levels did not differ of in the wild-type between the sufficient and deficient phosphate conditions, but increased in the SL-deficient mutants under phosphate deficiency, leading in the strong promotion of leaf senescence by GR24 treatment. These results indicate that the mutants exhibited increased responsiveness to GR24 under phosphate deficiency. In addition, GR24 accelerated leaf senescence in the intact SL-deficient mutants under phosphate deficiency as well as dark-induced leaf senescence. The effects of GR24 were stronger in d10 compared to d17. Based on these results, we suggest that SLs regulate leaf senescence in response to phosphate deficiency.

Article	Year
Tropane alkaloids in adventitious root cultures of Physochlaina physaloides. Kokuritsu Iyakuhin Shokuhin Eisei Kenkyujo hokoku = Bulletin of National Institute of Health Sciences\| PubMed ID: 12638186	2002
Tropane alkaloids in auxin-independent root cultures of Physochlaina physaloides. Kokuritsu Iyakuhin Shokuhin Eisei Kenkyujo hokoku = Bulletin of National Institute of Health Sciences\| PubMed ID: 12638187	2002
[Influence of fertilizers on yield and yield components of opium from Papaver somniferum Linn]. Kokuritsu Iyakuhin Shokuhin Eisei Kenkyujo hokoku = Bulletin of National Institute of Health Sciences\| PubMed ID: 14740399	2003
Ethylene induced shikonin biosynthesis in shoot culture of Lithospermum erythrorhizon. Plant physiology and biochemistry : PPB\| PubMed ID: 15820656	2005
Characteristics of Atropa belladonna hairy roots cryopreserved by vitrification method. Cryo letters\| PubMed ID: 16794738	2006
Co-expression of GbMYB1 and GbMYC1 induces anthocyanin accumulation in roots of cultured Gynura bicolor DC. plantlet on methyl jasmonate treatment. Plant physiology and biochemistry : PPB\| PubMed ID: 21123079	2011
Kinetic changes in glucosinolate-derived volatiles by heat-treatment and myrosinase activity in nakajimana (Brassica rapa L. cv. nakajimana). Journal of agricultural and food chemistry\| PubMed ID: 21913666	2011
Strigolactone signaling regulates rice leaf senescence in response to a phosphate deficiency. Planta\| PubMed ID: 24888863	2014

Koichiro Shimomura

Biography

JoVE Journal Publications

Other Publications