Genetička transformacija ukrasne koprive, Coleus blumei Benth. s pomoću bakterija Agrobacterium

Efficiency in transformation of three hybrids of Coleus blumei using wild and mutant strains of Agrobacterium sp. was evaluated. Successful transformation depended on specific bacterial strain-plant genotype combination and co-cultivation treatment. The wild type B6S3 of the bacterial strains employed gave the maximum efficiency. Strains C58C1(pArA4abc), GV3101(pGV2215), 8196 and A281 were also effective. Among several co-cultivation conditions tested, the most efficient treatment was as follows: excised leaf explants were incubated immeadiately in bacterial suspension for 5 min and were co-cultured with Nicotiana tabacum crown gall callus for two days. Plant hormone autotrophy, PCR analyses and hybridization analysis confirmed genetic transformation of the cultures. Sixteen different transgenic callus and cell cultures were maintained for more than four years in the absence of exogenous growth regulators and antibiotics without deterioration in the growth rate.
Tri genotipa ukrasne koprive (Coleus blumei Benth.) transformirana su s pomoću bakterija Agrobacterium tumefaciens i A. rhizogenes. Uspješnost transformacije ovisila je o sukultivacijskom postupku lisnih eksplantata i bakterija te o interakciji bakterijskoga soja i biljnoga genotipa. Soj divljeg tipa B6S3 bio je najvirulentniji. Najuspješniji je postupak transformacije bio petminutna inkubacija svježe narezanih lisnih eksplantata s bakterijama te nakon toga dvodnevna sukultivacija s tumorskim tkivom duhana (Nicotiana tabacum). Rast kalusnoga tkiva na hranidbenoj podlozi bez regulatora rasta, PCR analiza te hibridizacijska analiza potvrdili su genetičku transformaciju tkiva. Ustanovljeno je 16 transgeničnih kalusnih linija koje su se međusobno razlikovale po fenotipu i stopi rasta. Fenotipske značajke pojedine linije nisu se mogle povezati s bakterijskim sojem upotrijebljenim u poticanju transformiranih linija niti s biljnim genotipom.