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Jurnal Bioteknologi & Biosains Indonesia (JBBI) Pub Date : 2019-07-10 DOI:10.29122/JBBI.V6I1.3341

Suwinda Fibriani, Inyana Dwi Agustien, Widhi Dyah Sawitri, Bambang Sugiharto

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引用次数: 1

摘要

摘要蔗糖磷酸合酶(SPS)是参与蔗糖生物合成的关键酶。在调控过程中，SPS活性由一种变构效应物葡萄糖-6-磷酸(G6P)调节，该变构效应物被认为具有结合SPS n端结构域的能力。为了了解n端在SPS调控中的作用，我们对SPS基因进行突变，缺失n端结构域(∆N-SPS)。将∆N-SPS转化为番茄植株，转化效率为5%。获得3株转基因番茄植株4.20、5.5.1和5.10，经PCR分析证实。通过酶分析对转基因番茄的表达进行了表征。结果表明，转基因植株中G6P的变构调节丧失，SPS活性比非转基因植株提高2倍。这表明n端结构域缺失的SPS能够在植物中积极表达。关键词:酶，遗传转化，n端结构域缺失，蔗糖磷酸合成酶，番茄Dalam regulasinya, aktifitas SPS dipengaruhi oleh alosterik efektor glukosa-6-fosfat (G6P) yang diduga dapat berikatan pada结构域n端SPS。Untuk mengetahui peran n端pada regulasi SPS, dilakukan mutasi SPS dengan penghilangan域n端(∆N-SPS)。Gen∆N-SPS diinsersi paada tanaman番茄千层转化基因(melalui transformasi genetick dengan efisiensi transformasi 5%)。Tiga tanaman转基因番茄(event4.20;5.5.1;(5.10) didapatkan dan terkonfirmasi melalal分析PCR阳性。白藜芦醇的三聚氰胺分析。Hasil menunjukkan bahwa tanaman tomgenik (N-SPS tidak dipengaruhi regulasi alosterik G6P dan aktifitas SPS 2 kali lipat lebih tinggi daripada tanaman bukan transgenik。n -端结构域，n -端结构域，n -端结构域，n -端结构域，n -端结构域。Kata Kunci: delesi结构域n端，酶，蔗糖磷酸合酶，番茄，转化基因

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TRANSFORMASI GENETIK DAN EKSPRESI MUTAN SUCROSE PHOSPHATE SYNTHASE PADA TANAMAN TOMAT

Genetic Transformation and Expression of Sucrose Phosphate Synthase Mutant in Tomato Plant ABSTRACTSucrose phosphate synthase (SPS) is a key enzyme responsible for sucrose biosynthesis. In its regulation, SPS activity is modulated by an allosteric effector glucose-6-phosphate (G6P) suggested to have an ability to bind SPS N-terminus domain. To understand the role of N-terminus in regulating SPS, the SPS gene was mutated with the deletion of N-terminus domain (∆N-SPS). The ∆N-SPS gen was transformed into tomato plants with 5% transformation efficiency. Three transgenic tomato plant 4.20, 5.5.1, and 5.10 were obtained and confirmed by PCR analysis. Transgenic tomato expression was characterized by enzymatic analysis. Result showed that the G6P allosteric regulation in transgenic ∆N-SPS had lost and the SPS activity increased by 2-fold compared to non-transgenic plant. This showed that N-terminus domain-deleted SPS could be actively expressed in plant. Keywords: enzyme, genetic transformation, N-terminus domain deletion, sucrose phosphate synthase, tomato ABSTRAKSucrose phosphate synthase (SPS) merupakan enzim kunci yang bertanggung jawab dalam sintesis sukrosa. Dalam regulasinya, aktifitas SPS dipengaruhi oleh alosterik efektor glukosa-6-fosfat (G6P) yang diduga dapat berikatan pada domain N-terminus SPS. Untuk mengetahui peran N-terminus pada regulasi SPS, dilakukan mutasi SPS dengan penghilangan domain N-terminus (∆N-SPS). Gen ∆N-SPS diinsersi pada tanaman tomat melalui transformasi genetik dengan efisiensi transformasi 5%. Tiga tanaman transgenik tomat (event4.20; 5.5.1; dan 5.10) didapatkan dan positif terkonfirmasi melalui analisis PCR. Ekspresi mutan dikarakterisasi melalui analisis enzimatik. Hasil menunjukkan bahwa tanaman tomat transgenik ∆N-SPS tidak dipengaruhi regulasi alosterik G6P dan aktifitas SPS 2 kali lipat lebih tinggi daripada tanaman bukan transgenik. Ini menunjukkan bahwa SPS dengan delesi domain N-terminus dapat terekspresi aktif pada tanaman. Kata Kunci: delesi domain N-terminus, enzim, sucrose phosphate synthase, tomat, transformasi genetik

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Jurnal Bioteknologi & Biosains Indonesia (JBBI)

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