Abstract
<p class="p1">The experiment was conducted at the Molecular Biology Laboratory of the Indonesian Center for Agricultural Biotechnology and Genetic Resources Research and Development, Bogor. The objective was to construct <em>-ai </em>gene on a binary plasmid <em>p</em>Cambia 1301. This experiment was carried out using construction method by ligation process between fragments of <em>α-ai </em>gene from <em>p</em>TA<span class="s1">3 </span>plasmid and <em>p</em>Cambia 1301 on <em>Hind</em>III site. The result of ligant transformation into <em>E. coli </em>DH5<em>α </em>was 182 surviving colonies on YEP medium containing kanamycin. DNA samples were obtained from 60 randomly selected colonies. The restriction pattern was tested by digesting each DNA sample using <em>Hind</em>III showed colonies containing two fragments expected of sizes wich are 11.837 and 4.887 kb. Two colonies are predicted containing of <em>α-ai </em>gene on its the binary plasmid. Advanced tests using restriction enzymes <em>Bam</em>HI and <em>Xba</em>I showed two directions (right and left) of <em>α-ai </em>gene. The right direction was shown by <em>p</em>Cambia-<em>α-ai</em>1 from colony number 43. This plasmid showed expected fragments of sizes 13.485 and 3.219 kb when digested with <em>Bam</em>HI and two fragments of sizes 15.421 and 1.303 kb when digested with <em>Xba</em>I. The left direction was shown <em>p</em>Cambia-<em>α-ai</em>2 from colony number 58. This plasmid also demon-strated expected fragments of sizes 15.026 and 1.698 kb when digested with <em>Bam</em>HI and two fragments of sizes 13.082 and 3.642 kb when digested with <em>Xba</em>I. Both <em>p</em>Cambia-<em>α-ai</em>1 and <em>p</em>Cambia-<em>α-ai</em>2 were transformed into <em>A. tumefaciens </em>LBA4404.</p>
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