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1 APPENDIX 1. hIURASRICE AND SI(SO) --.- I 0.025 ~- ~ OR Orgn~iics (X100) 27 8 2.78 giL Glycine 200 mdL ~ 10 1111 h'icotir~ic acid ) 10 1111 Thiamine HCI Pyridoxine HCI i 0.5 100 myL 111-lnositol ! 100 5.0 g5UU rill I I0 1i1i
2. Fixative used for fisntio~~ of tissue culture sun~l~lcs: Acetic acid and 95% Etllallol were ~ilixed at 1:3 proportions added. The co~ripo~ie~its wcrc sliakeri tint11 ill1 oftllelir ;~rc ill sol~~tlo~l. 'l'llu color oitl~c stain was redd~sll violct. Fol. Eosin stock sulutioli, I y of liosili (w:iIci. solilblc) \\as dissolved in 20 1111 ofdistilled water and 111;ldc lip to IUU rill \\it11 05'i." ctl~;~~lol 3. DNA exlrilctio~~ buffer (DELLAPOR'SA 111et11od) 100 nlM Tris pH8 4. Pri~nilry IV:I~~I buffer (I litre) (for Soutl~er~~ i~n:llysis) 1 SDS 0.5% 1 5. Secondary a'asl~ buffer (ZOS stock)(for Suutl~err~ c~~l:tlysis) Tris base I21 g. IM I'laCI 1 12 g. ?M Adjust pH to 10.0. M3ix i~pto 1 Iilrc wit11 water, l'h~s call be kept for up to 4 months. 1 :20 I.e., 2 mlIL of lbi MsCl>
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GENETIC TRANSFORMATION OF Cicer ari
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Certificate Ger/$ed /La/ /Ae eldire
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I deem il as a great pleastire to e
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CONTENTS ABBREVIAI'IONS SUMMARY LIS
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3.2.2.3 Shoot elongat~on 3.2.2.4 Ro
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ABBREVIATIONS 2-il' 2-isopci~~cnpl
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frequency was observed wit11 2,4,5-
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in vitro grown plants w;is ocli~eve
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LIST OF TABLES Table 3.1 Pr~~iier c
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Table 4.1 1 Effect ul'age uf lhe ex
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Figure 4.1 A - D inducl~on of sonla
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Figure 4.9 A - C Alternative llleti
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Figure 4.15 A, B Restriction analys
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Figure 4.20 A, B DNA profile of gen
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1'1.0 INTRODUCTION L~fe is the ter~
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them. About 10% of the Earth's land
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observations GkI Shull's experiment
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(Nene and Haware, 1980). The legume
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jf.0 REVIEW OF LITERATURE Large par
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plants from them enlhuscd scientist
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introduction into protoplasts using
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evident by 1983 that the Agrobacter
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The above promoters were found to b
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gained an illstant popi~larity owi~
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legume crop species. Cotyledon expl
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1994). Clonal propagation of FI int
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were induced to for111 callus when
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' the ernblyos into plantlets on MS
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and nprII for selection. PCR and So
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Agrobacleriuiii and ill otlier expe
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ioteclinologicsl i~iiprovemel~t of
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combinat~ons and concentrdtions of
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egenerated via somatic eliibryogene
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egenerat~on from protoplasts and ca
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transformants were identified by sy
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urgent econoniical control measures
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applications of traps. Technically
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in the laboratory and releasing tl~
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have been exhaustively reviewd (Dai
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larvae of certaln Insects were unab
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3.1 Pl:~nt 111nteri:il sllrl cultur
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0.5. 2.0 and 5.0 pM were added. 111
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Lec!/i?is: Young anci juve~iile lea
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AM3 explant: This was a by-product
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PM) 2nd GAJ (2 5 PM), eltller singl
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of the plastic bag was removed rwlc
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- . -- laopropyl alcoliol ; Absolul
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The sl~des were Illell muiaited and
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6. The bupcniatant \\,I,, t,ikcli i
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5. Microcal~lers were allowed to se
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till the growtli of bacterial cells
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;I! 37 'C for 3-24 Iiours 111 d;irL
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PCN for 111cl.4 ge!iiJ: I'C'I< tbr
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4. Tliu pel \\as \r;~alicd 3 ti~iir
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4.0 RESULTS $loaf of tllc c\l~cr~~l
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These combi~iatiori sl~owed liiorc
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egeneratloll \!;IS dli'licull ;~nti
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decreased a[ cllkalinc pH \+li~le 1
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either side of'bws;tl portiol~ oill
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IBA sl~owed bro\\~~i~~g i~lld de:~l
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humidity uf 50% 111 the dayrinie a1
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4.2 Histological studies ~II nlultl
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sliowetl no liirtlie~ yrowlli Tllc
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Conlirmntioll of plllati!e t~.~ll~a
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5.0 DISCUSSION During tile la31 dcc
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leatlets (Dilieshkuniar er al.. 109
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used with an add111ou;ll care :u~d
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tissue to regenerate illto lesser i
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prololigcd culture on TUL sho\\~d 1
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exceed 5 cni Icngtl~. Loiigcr shoot
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stem and lravca Secol~d;lrq ~'IIICI
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arnplitication of the sopccIi\i. 90
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Explant Preparation -Geminate seeds
Page 150 and 151: 6.0 REFERENCES Adkins, A.L.. Godwin
Page 152 and 153: Avenido, R.A., Hattori, I
Page 154 and 155: Benfey, P.N., Chua, N.1-I., Ilcgiii
Page 156 and 157: Cnrvalho. IC1.H C dc , Uui Vat1 LC
Page 158 and 159: Chu, P.W.G., Andersoli. B.J., Khan.
Page 160 and 161: Deblaere, R., Reyneerts, A., Hotte,
Page 162 and 163: Drun~mond, M.H., Gordoil. M.P., Nes
Page 164 and 165: Foster, J.G., Coulombe, B.A., Van S
Page 166 and 167: Gill, R., Saxstla, I'.K., (1992) Di
Page 168 and 169: Hess, D., (1970) Versueche zur tran
Page 170 and 171: Hopkinson, M.J. ;md Collins, I1.M.,
Page 172 and 173: Karthikeyan, A.S., Sarina, K.S., Ve
Page 174 and 175: Krens, F.A., Molendijk, L., L., Wul
Page 176 and 177: Li, Z.Y., Tanner, G.J., Larkin. P.J
Page 178 and 179: Maxan~, A.M., Gilbert, W., (1977) A
Page 180 and 181: field conditions. Procc.rdbigs o/Na
Page 182 and 183: Ohara, A., Akasako, Y., Daimon, H.,
Page 184 and 185: llechrnlics sur les prnidilies des
Page 186 and 187: Rand, G.M. (Ed.) (1995) Ftordir~~rc
Page 188 and 189: Richardson, M.J., (199 I ) Seed sto
Page 190 and 191: Sarria, R., Calderon, A., Thro, A.M
Page 192 and 193: Shri, P.V., Davls, T.M., (1992) Zen
Page 194 and 195: Tegeder, M. Koli~i, I-I., Nibbbe, M
Page 196 and 197: Venkatachalam, P., Klshor, P.B.K.,
Page 198 and 199: Winans, S.C., (1992) Tiyo way chemi
Page 203 and 204: Table 4.1 Induction of sorrlalic er
Page 205 and 206: Table 4.3 111ducti011 of en~bryos O
Page 208 and 209: 'Table 4.6 Dificrent combinations o
Page 210 and 211: Table 4.8 Effect oI'TDZ, 2-iP :~nd
Page 212 and 213: Table 4.10 Effect of i~lclusio~~ or
Page 214 and 215: Table 4.12 Induction of multiple sh
Page 216 and 217: Table 4.14 Effect of media constitu
Page 218: Figure 3.1 Dii~gralll~l~i~tic rcpre
Page 222: Figure 3.3 A - C Diagrn~nn~atic rep
Page 225 and 226: I . Nhu 1 51 ?C,%- .pal5125 &pa1512
Page 228: I
Page 232: Induction of somatic embryos from m
Page 238: I.iy111. J.5 SI:i;i.\ iil ~ ~ l ~ ~
Page 254:
IIistologic:~l stutlic\ of dcveiopr
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Rc\[ricfiot~ ;111;1l)si\ ur ill? ~
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I'C'R :~llipLiiir:iriuil 01'700 bl)
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Fixi1r.c 17 \r:15 -l)K,.\ ;!lid J"
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1:igurr 4.22 Sui~lllcrn :III:II!s~$
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