Citokininska homeostaza i ekspresija gena uključenih u kontrolu ćelijskog ciklusa tokom procesa regeneracije kelerabe (Brassica oleracea L. var. gongylodes) in vitro
Cytokinin homeostasis and expression of the genes involved in cell cycle control during kohlrabi (Brassica oleracea L. var. gongylodes) regeneration in vitro
Doktorand
Ćosić, Tatjana M.Mentor
Ninković, SlavicaČlanovi komisije
Dragićević, IvanaRadović, Svetlana
Metapodaci
Prikaz svih podataka o disertacijiSažetak
Regeneracija in vitro gajenog biljnog tkiva kontrolisana je regulatorima rastenja,
naročito citokininima i auksinima. Egzogeno primenjeni biljni regulatori rastenja mogu
interagovati sa endogenim biljnim hormonima i uticati na njihovu biosintezu ili
distribuciju, odnosno menjati hormonalnu homeostazu koja utiče na razviće in vitro.
Proces regeneracije biljaka in vitro podrazumeva intenzivne ćelijske deobe, a centralnu
ulogu u kontroli ćelijskog ciklusa i razvića organa imaju kompleksi ciklina i kinaza
zavisnih od ciklina na čiju sintezu, odnosno aktivnost, utiču pomenuti biljni hormoni. U
tom smislu, ciljevi ovog rada bili su uspostavljanje efikasnih protokola za regeneraciju
biljaka kelerabe (Brassica oleracea L. var. gongylodes) in vitro, sa posebnim osvrtom
na identifikaciju i kvantifikaciju različitih grupa citokinina i indol-3-sirćetne kiseline
(IAA) u eksplantatskim tkivima sa regenerisanim izdancima, kao i analiziranje obrazaca
i intenziteta ekspresije gena koji kodiraju cikline... i kinaze zavisne od ciklina tokom de
novo organogeneze izdanaka. Efikasan sistem za repetitivnu somatsku embriogenezu
(SE) kod kelerabe, sorta Bečka Plava (BP), je uspostavljen iz nezrelih zigotskih
embriona. Najveća frekvencija SE detektovana je kod nezrelih zigotskih embriona u
ranom kotiledonarnom stadijumu razvića gajenih na hranljivoj podlozi bez regulatora
rastenja.
Uspešna indukcija de novo organogeneze izdanaka kelerabe, sorte BP i Bečka Bela
(BB), postignuta je kod odsečaka hipokotila i intaktnih klijanaca, na podlogama sa
pojedinačnim citokininima (CK): 6-benzilaminopurin (BAP), tidiazuron (TDZ), transzeatin
(transZ) ili cis-zeatin (cisZ). Frekvencija regeneracije kao i vijabilnost
regenerisanih izdanaka obe sorte je bila veća kod intaktnih klijanaca, dok su među
primenjenim citokininima BAP i TDZ bili najefikasniji u indukciji de novo
organogeneze. Kvantifikacija endogenog sadržaja citokinina i IAA u odsečcima
hipokotila sa regenerisanim izdancima (HRI) kao i u intaktnim klijancima sa
regenerisanim izdancima (KRI) je ukazala na to da uočene razlike u organogenom
odgovoru ova dva tipa početnog biljnog materijala mogu biti povezane sa njihovim
hormonskim statusom...
vitro plant regeneration is under control of plant growth regulators (PGRs), especially
cytokinins and auxins. Exogenously applied PGRs can affect the homeostasis of
endogenous plant hormones by influencing their biosynthesis and distribution,
subsequently altering in vitro development. Intensive cell divisions represent the basis
of in vitro plant regeneration, and they are controlled by complexes of cyclins and
cyclin-dependent kinases. The activity of these complexes during cell cycle and organ
development is affected by plant hormones. Respecting this, the aims of this study were
developing efficient protocols for in vitro regeneration of kohlrabi (Brassica oleracea
L. var. gongylodes) with an emphasis on identification and quantification of different
groups of endogenous cytokinins and indole-3-acetic acid (IAA) in explant tissue with
respective regenerated shoots, as well as analyzing expression of genes encoding cyclins
and and cyclin-dependent kinases during de novo shoot organ...ogenesis. Efficient system
for repetitive somatic embryogenesis (SE) was developed for kohlrabi cv. Vienna
Purple (VP) using immature zygotic embryos. The highest frequency of SE was
obtained from immature zygotic embryos at the early cotyledonary stage grown on
PGR-free medium.
Successful induction of de novo shoot organogenesis in kohlrabi cv VP and cv. Vienna
White (VW) was achived using hypocotyl explants and intact seedlings, cultivated on
media supplemented with single cytokinin (CK): 6-benzylaminopurine (BAP),
thidiazuron (TDZ), trans-zeatin (transZ) or cis-zeatin (cisZ). Higher frequency of shoot
regeneration as well as viability of regenerated shoots was detected for intact seedlings,
with BAP and TDZ being the most efficient in de novo organogenesis induction.
Quantification of endogenous cytokinin and IAA contents in both hypocotyl explants
and seedlings with regenerated shoots (HRSs and SRSs, respectively) suggested that the
observed differences in organogenic response between these two types of starting plant
material were related to their cytokinin and IAA contents. The level of total CKs was
higher in HRSs, while SRSs displayed a higher IAA/bioactive CKs ratio...