Study of in vitro selection and plant regeneration of Indica rice tolerant to iron

AȱandȱB).ȱTheȱmaximumȱproliferationȱofȱembryogenicȱ andȱotherȱculturesȱdidȱnotȱshowȱanyȱsignȱofȱgrowthȱ

callusesȱ isȱ notedȱ inȱ theȱ mediumȱ containingȱ 1.0ȱ andȱ turnedȱ brownȱ withinȱ oneȱ weekȱ ofȱ culture.ȱ

mg/lȱ Knȱ supplementedȱ withȱ 3.0ȱ mg/lȱ 2,4-D.ȱ BAȱ isȱ Subsequently,ȱ theȱ callusesȱ culturedȱ onȱ 10ȱ andȱ 12ȱ

notȱasȱeffectiveȱinȱembryogenicȱcallusȱinductionȱasȱ mMȱ grewȱ rapidlyȱ intoȱ yellowish-whiteȱ colorationȱ

comparedȱ withȱ Kn.ȱ Theȱ mediumȱ havingȱ Knȱ withȱ (Figureȱ1C).ȱTheȱculturesȱthatȱshowedȱgrowthȱwereȱ

2,4-Dȱwasȱobservedȱtoȱbeȱbestȱforȱcallusȱproliferation.ȱ separatedȱandȱsubculturedȱonȱrespectiveȱmediumȱatȱ

Theȱ proliferatedȱ primaryȱ globularȱ callusesȱ areȱ fourȱweekȱintervals.ȱTheȱtolerantȱcallusesȱareȱagainȱ

subsequentlyȱ subculturedȱ intoȱ similarȱ mediumȱ transferredȱontoȱaȱsimilarȱmediumȱwithȱ12ȱmMȱFeȱ

forȱ furtherȱ proliferationȱ ofȱ embryogenicȱ calli.ȱ Theȱ orȱ controlȱ medium.ȱ Significantȱ growthȱ isȱ achievedȱ

mediumȱdevoidȱofȱgrowthȱregulatorsȱdidȱnotȱhelpȱ onȱ mediumȱ containingȱ 12ȱ mMȱ Feȱ (tolerant)ȱ asȱ

inȱ theȱ proliferationȱ ofȱ callusȱ development.ȱ Theȱ comparedȱwithȱtheȱcontrolȱmediumȱ(non-tolerantȱ).ȱ

maximumȱ rateȱ ofȱ embryogenicȱ callusȱ proliferationȱ TheȱGTIȱofȱnon-tolerantȱandȱtolerantȱcallusȱshowedȱ

wasȱ notedȱ inȱ MSȱ mediumȱ supplementedȱ withȱ significantȱvariationȱonȱmediumȱcontainingȱdifferentȱ

0.5–1.0ȱ mg/lȱ Kn,ȱ 3.0ȱ mg/lȱ 2,4-Dȱ inȱ O.sativa Vars.

concentrationsȱofȱFeȱ(Tableȱ2).ȱTheȱtolerantȱcallusesȱ

Khandagiriȱ andȱ Jagannath.ȱ Theȱ proliferationȱ ofȱ producedȱsignificantlyȱmoreȱfreshȱandȱdryȱbiomassȱatȱ

friableȱembryogenicȱcallusȱinȱtermsȱofȱfreshȱweightȱ 12ȱmMȱFeȱthanȱtheȱnon-tolerantȱcultures.ȱTheȱgrowthȱ

wasȱ maximumȱ inȱ theȱ mediumȱ havingȱ Knȱ +ȱ 2,4-

ofȱ non-tolerantȱ callusesȱ is,ȱ however,ȱ stimulatedȱ byȱ

Dȱ asȱ comparedȱ toȱ Knȱ orȱ BAȱ +ȱ NAAȱ andȱ showedȱ theȱlowȱconcentrationȱofȱFe.ȱWithȱfurtherȱincreasesȱ

yellowishȱwhiteȱcoloration.ȱSimilarȱresponsesȱwereȱ inȱFeȱconcentration,ȱtheȱcallusȱshowedȱaȱreductionȱ

observedȱ whenȱ 2,4-Dȱ isȱ replacedȱ withȱ NAA.ȱ Theȱ inȱ freshȱ weightȱ byȱ 40%ȱ atȱ 12ȱ mMȱ Fe.ȱ Theȱ growthȱ

embryogenicȱ calliȱ developmentȱ inȱ somaticȱ cellsȱ isȱ isȱ arrestedȱ dueȱ toȱ theȱ highȱ concentrationȱ ofȱ Feȱ inȱ

oftenȱaccompaniedȱcellularȱstress.ȱMoreover,ȱ2,4-D,ȱ theȱ growthȱ media,ȱ whichȱ mayȱ beȱ bindingȱ proteinȱ

theȱ mostȱ frequentlyȱ usedȱ forȱ inductionȱ ofȱ somaticȱ (Steffens,1990).ȱ Theȱ tolerantȱ andȱ non-tolerantȱ

embryogenesis,ȱwasȱknownȱtoȱactivateȱmanyȱstressȱ callusesȱ areȱ maintainedȱ forȱ aȱ prolongedȱ periodȱ onȱ

relatedȱgenesȱsupportingȱtheȱhypothesisȱthatȱsomaticȱ similarȱmedium.

embryogenesisȱ resultedȱ dueȱ toȱ extremeȱ stressȱ

responseȱofȱculturedȱcells.ȱInȱsomeȱcases,ȱhowever,ȱ Pigment and Protein Analysis

inductionȱmediumȱfavoursȱbothȱcallusȱinductionȱandȱ

Bothȱ tolerantȱ andȱ non-tolerantȱ callusesȱ areȱ takenȱ

stimulationȱofȱembryogenicȱresponse.ȱTheȱabilityȱtoȱ

forȱ biochemicalȱ analysis.ȱ Theȱ callusȱ derivedȱ fromȱ

formȱ embryogenicȱ callusȱ isȱ genotypeȱ dependent.ȱ

tolerantȱlinesȱshowedȱtheȱhigherȱchlorophyllȱcontentȱ

Outȱofȱtwoȱgenotypesȱtested,ȱoneȱgenotypeȱfavoredȱ

asȱcomparedȱwithȱnon-tolerantȱlinesȱ(Tableȱ3).ȱSimilarȱ

maximumȱ embryogenicȱ frequencyȱ asȱ comparedȱ toȱ

observationsȱ wereȱ madeȱ earlierȱ onȱ chlorophyllȱ

anotherȱgenotype.

synthesisȱ inȱ tobaccoȱ callusȱ dueȱ toȱ theȱ presenceȱ ofȱ

theȱhighȱconcentrationȱofȱmanganeseȱ(Petolinoȱandȱ

Selection of Fe tolerant Embryogenic Callus

Collinsȱ1985;ȱClairmont et al. 1986).ȱTheȱcholophyllȱaȱ

EmbryogenicȱcalliȱgrownȱonȱmediumȱcontainingȱMSȱ andȱbȱcontentȱinȱtheȱtolerantȱsourcesȱincreasedȱ58.77ȱ

basalȱsaltsȱsupplementedȱwithȱ1.0ȱmg/lȱKnȱandȱ3.0ȱ toȱ117.8%ȱforȱchlorophyllȱaȱandȱ68.07ȱtoȱ81.63%ȱforȱ

mg/lȱ2,4-Dȱwereȱtransferredȱtoȱsubcultureȱmediumȱ chlorophyllȱ bȱ inȱ theȱ presenceȱ ofȱ 12ȱ mMȱ Feȱ (Tableȱ

havingȱsimilarȱcompositionȱbutȱsupplementedȱwithȱ 3).ȱ Theȱ proteinȱ contentȱ isȱ moreȱ inȱ tolerantȱ calliȱ asȱ

differentȱconcentrationsȱofȱFeȱ(1.5ȱmMȱ(Control),ȱ4,ȱ comparedȱwithȱnon-tolerantȱcalliȱ(Tableȱ4).ȱMandal et

8,ȱ10,ȱ12ȱandȱ14ȱmM).ȱTheȱcallusȱrapidlyȱproliferatedȱ al. (2004)ȱreportedȱthatȱtheȱFe-tolerantȱriceȱcultivarsȱ

intoȱprofuseȱyellowishȱwhiteȱstructuresȱinȱ4ȱweeksȱ absorbȱ lessȱ Feȱ fromȱ rootȱ toȱ leaves,ȱ indicatingȱ theȱ

onȱ mediumȱ containingȱ 4ȱ andȱ 8mM.ȱ Fewȱ coloniesȱ governanceȱofȱaȱphysiologicalȱavoidanceȱmechanismȱ

grewȱatȱhigherȱconcentrationsȱofȱ10ȱandȱ12ȱmMȱFeȱ atȱanyȱconcentrationȱofȱFeȱinȱtheȱleaves.ȱFe-tolerantȱ

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