hrvatski

Razvojno specifična ekspresija proteina u kulturi biljnoga tkiva

Nakon nekoliko desetljeća intenzivnih istraživanja i važnih otkrića o strukturi i funkciji gena, suočeni smo danas sa sve većim zanimanjem za proteom – funkcionalni komplement genoma. Eksperimantalni pristup obično uključuje dvodimenzionalnu elektroforezu, nakon koje može slijediti identifikacija proteina masenom spektrometrijom. U ovome radu istraživalo se biljni razvitak i morfogenezu u kulturi in vitro i to na razini uzorka izraženih proteina. Istraživalo se na modelu triju biljnih vrsta: šećerne repe (Beta vulgaris L.), hrena (Armoracia lapathifolia Gilib.) i kaktusa (Mamillaria gracillis Pfeiff.). Razvojni procesi potaknuti su unutarnjim staničnim čimbenicima, okolišnim uvjetima ili transformacijom stanica plazmidom Ti (divlji tip B6S3), a bez djelovanja vanjskih regulatora rasta. Otkrivene su promjene u programima aktivacije gena na razini staničnih i izvanstaničnih proteina i izoenzima (peroksidaze, esteraze i invertaze). Proteini su analizirani jednodimenzionalnom i dvodimenzionalnom elektroforezom, obojeni srebrom ili su karakterizirani lektinima (Con A, UEA, RCA120). Razvojno specifični uzorci proteinskih mrlja dobiveni dvodimenzionalnom elektroforezom omogućit će identifikaciju nekih tkivno-specifičnih proteina masenom spektrometrijom. After many decades of intensive genome research and impressive revelations on gene structure and function we are facing the growing interest on a proteom - a functional genome complement. The experimental approach usually includes two-dimensional gel electrophoresis, which can be combined with protein identification by mass spectrometry. In the present work plant development and morphogenesis in in vitro culture were studied at the level of protein expression pattern. Three model species: sugar beet (Beta vulgaris L.), horseradish (Armoracia lapathifolia Gilib.) and cactus (Mamillaria gracillis Pfeiff.) have been studied. Developmental processes were triggered by intrinsic cellular factors, environmental condition and by Ti-plasmid (a wild type B6S3) transformation, without the action of any exogenous growth regulators. Changes in gene activation programs were detected at the level of cellular and extracellular proteins and isoenzymes (peroxidases, esterases, invertases). Proteins were analysed by means of one-dimensional and two-dimensional electrophoresis, silver-stained or characterised by lectins (Con A, UEA, RCA120). Developmentally specific protein patterns obtained by means of 2-D electrophoresis should enable us to identify some of tissue specific proteins by means of mass spectrometry.

kultura biljnoga tkiva

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