New Researches in Biotechnology - Facultatea de Biotehnologii ...

More documents

Recommendations

Info

Proceeding of the 4 rd International Symposium“NEW RESEARCH IN BIOTECHNOLOGY” USAMV Bucharest, Romania, 2011plants, therefore the use virus-free vines for multiplication and new vineyard establishingis required (F. MANNINI [1]). In cases of high incidence of infectious diseases in breedingvineyards and impossibility to select authentic and disease-free material of a particularvariety or rootstock, several techniques are being applied for virus elimination (I.TSVETKOV & al. [ 2]).The meristem culture and thermotherapy are individually or in combination themost frequent methods of obtaining virus-free grapevines (G. GRAMMATIKAKI & A.AVGELIS [3]; B. KŘIŽAN & al. [4]) The application of heat treatment for viruseliminationin plants is time and energy consuming and needs special equipments(chambers for heat treatment). In the case of the meristem culture, the percent of viruselimination is influenced by the genotype, the type of the virus and especially of thedifficulty of meristem excision.Virus-elimination techniques such as: chemotherapy (A. PANATTONI & al.[5]), electrotherapy in continuous electric field (I.C. GUȚĂ & al. [6]), cryotherapy (SH.BAYATI & al. [7]), somatic embryogenesis (G. GAMBINO & al. [8]) and theircombinations are considered alternative method to thermotherapy. Each of the listedmethod involves one stage of in vitro culture.Is well known the use of electric current in the cleaning of viral diseases in plants;instead, a deep knowledge of this phenomenon theoretical basis is not yet available. Theexperiments demonstrated that the cleaning process nature is, essentially, the effect of theviral protein denaturation, by means of the heat to which they were exposed in the thermalbath that the vegetal tissue constitutes (J.E GONZÁLES & al. [9]).Electrotherapy has been applied successfully to eradicate potato virus X fromdifferent infected potato clones (H.F. LOZOYA-SALDAÑA & al. [10]); dasheen mosaicvirus in malanga (J. IGARZA CASTRO & al. [11]; banana streak virus in banana (P.R.HERNÁNDEZ & al. [12]).The electrotherapy for GFLV elimination was investigated, as a more practicalalternative technique comparatively to heat therapy of virus infected grapevine, but notsatisfactory results have been obtained. However, the decreasing of by enzyme-linkedimmunosorbent assay (ELISA) values in function of the exposure period have beenregistered (J.G. BURGER [13]).The electrotherapy followed by in vitro culture of shoot apices produced 33.33–66.66% GLRaV-1+3 –free plants, regardless explant position of exposed grapevine incontinuous electric field (I.C. GUȚĂ & al. [14]).The aim of the present research was to eliminate fanleaf virus (GFLV), arabismosaic virus (ArMV), leafroll associated virus 1 (GLRaV-1), leafroll associated virus 342
Proceeding of the 4 rd International Symposium“NEW RESEARCH IN BIOTECHNOLOGY” USAMV Bucharest, Romania, 2011(GLRaV-3), and fleck virus (GFkV) in simple or mixed infections at a time, in seven Vitisvinifera L. varieties, by electrotherapy with direct electric current.2. MATERIALS AND METHODSSource of virus-infected material: The study regarding GFLV, ArMV, GLRaV-1,GLRaV-3, GFkV and their combinations (virus complex) elimination has been done onseven V. vinifera L. cultivars naturally infected (Table 1), maintained in the grapevinevirus infected collection of NRDIBH Ştefăneşti–Argeş. One year old infected plantsobtained from one bud cuttings, confirmed as virus-infected by ELISA testing, constitutedthe source of nodal fragments (microshoots).Table 1. Virus-infected grapevine cultivars used in the experimentGrapevine cultivarVirus/virus complexFetească albăGFLVFetească neagrăGLRaV-1GLRaV-1FrâncușăCabernet SauvignonGrasă de CotnariUnidentified genotype (1)Unidentified genotype (2)GLRaV-3GFkVArMV + GFkVGFLV + GLRaV-1Electrotherapy: In the case of electrotherapy with direct electric current, a sourcethat converts the alternating current from electricity network in alternating current withadjustable parameter is required. The experiment consists of subjecting a nodal fragment toan alternating current for a certain period, after that it is inoculated on culture medium,being the source of regeneration of new grapevine plants.The experimental parameter were established after previous attempts (directelectric current of 1; 100; 1000 and 10 000 kHz was applied for 5; 10 and 20 min at thecut ends of one budded herbaceous cuttings), regarding the viability of microshootsexposed to the action of electric current (unpublished data). The current frequency (1000kHz; 10 000 kHz) and exposure time (10 min) for this experiment were established. Thus,the experience is bifactorial of subdivided parcels type, 2x7 experimental variants; for eachvariant were exposed three microshoots as repetitions.The control for each variant was represented by virus infected microshoots in vitrocultured unexposed to direct electric curent (untreated). The position of the nodal fragmentson the shoots were identically for all experimental variants, namely the second internodebelow the apex.In vitro culture: Nodal fragments subjected to electrical current treatment andvirus- infected untreated nodat fragments (control) have been inoculated on M&S (1962)basic medium (T. MURASHIGE & F. SKOOG [15]) containing 2 µmol L -1benzylaminopurine (BAP) and 5 µmol L -1 indolilacetic acid (AIA). After three subcultures43
Page 1 and 2: UNIVERSITY OF AGRONOMICAL SCIENCES
Page 3 and 4: Proceeding of the 4 rd Internationa
Page 41: Proceeding of the 4 rd Internationa
Page 93 and 94:
Proceeding of the 4 rd Internationa
Page 95 and 96:
Page 97 and 98:
Page 99 and 100:
Page 101 and 102:
Page 103 and 104:
Page 105 and 106:
Page 107 and 108:
Page 109 and 110:
Page 111 and 112:
Page 113 and 114:
Page 115 and 116:
Page 117 and 118:
Page 119 and 120:
Page 121 and 122:
Page 123 and 124:
Page 125 and 126:
Page 127 and 128:
Page 129 and 130:
Page 131 and 132:
Page 133 and 134:
Page 135 and 136:
Page 137 and 138:
Page 139 and 140:
Page 141 and 142:
Page 143 and 144:
Page 145 and 146:
Page 147 and 148:
Page 149 and 150:
Page 151 and 152:
Page 153 and 154:
Page 155 and 156:
Page 157 and 158:
Page 159 and 160:
Page 161 and 162:
Page 163 and 164:
Day 7Day 10Proceeding of the 4 rd I
Page 165 and 166:
Page 167 and 168:
Page 169 and 170:
Page 171 and 172:
Page 173 and 174:
Page 175 and 176:
Page 177 and 178:
Page 179 and 180:
Page 181 and 182:
Page 183 and 184:
Page 185 and 186:
Page 187 and 188:
Page 189 and 190:
Page 191 and 192:
Page 193 and 194:
Page 195 and 196:
Page 197 and 198:
Page 199 and 200:
Page 201 and 202:
Page 203 and 204:
Page 205 and 206:
Page 207 and 208:
Page 209 and 210:
Page 211 and 212:
Page 213 and 214:
Page 215 and 216:
Page 217 and 218:
Page 219 and 220:
Page 221 and 222:
Page 223 and 224:
Page 225 and 226:
Page 227 and 228:
Page 229 and 230:
Page 231 and 232:
Page 233 and 234:
Page 235 and 236:
show all

New Researches in Biotechnology - Facultatea de Biotehnologii ...

Create successful ePaper yourself

Delete template?

Save as template?