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RISS 인기검색어
- 검색키워드
- 저자 : M. Aydın Akbudak (검색결과 3 건)
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Suppression of Arabidopsis genes by terminator-less transgene constructs
M. Aydın Akbudak,Scott J. Nicholson,Vibha Srivastava 한국식물생명공학회 2013 Plant biotechnology reports Vol.7 No.4
Transgene-mediated gene silencing is an important biotechnological and research tool. There are several RNAi-mediated techniques available for silencing genes in plants. The basis of all these techniques is to generate double-stranded RNA precursors in the cell, which are recognized by the cellular surveillance system, and marked for degradation by the Dicer family RNases into siRNAs. Improperly terminated, unpolyaden lated RNA are precursors of double-stranded RNA, and, therefore, can serve as silencing triggers in plants. Such transcripts can easily be synthesized from transgene constructs lacking transcription-terminator signals (terminator-less constructs). The present study determined the silencing efficiency of terminator-less constructs on six different genes in Arabidopsis: Phytochrome A (PHYA), Brassinosteroid Insensitive 1 (BRI1), Variegated 2 (VAR2), Constans (CO), Apetala 1 (AP1) and Transparent TestaGlabra 1 (TTG1). Expression of terminator-less gene fragments of PHYA, AP1, and VAR2 resulted in a *90 % decline, and those of BRI1 and CO resulted in a *70 % decline, in the steady state level of the respective transcript in transgenic lines compared to the wild-type. This suppression was accompanied by phenotypic aberrations inselected transgenic lines. Thus, targeted gene suppression in plants can be initiated by the expression of a simple construct design consisting of a gene fragment lackingtranscription terminator signals
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Sathish K. Ponniah,Zhenhua Shang,M. Aydın Akbudak,Vibha Srivastava,Muthusamy Manoharan 한국식물생명공학회 2017 Plant biotechnology reports Vol.11 No.1
Rice straw is one of the largest biomasses in the world that can potentially be exploited for biofuel. Nevertheless, the association of lignin with cellulose and hemicellulose has hindered the efficient utilization of rice straw for cellulosic biofuel. The objective of this study was to down-regulate key genes involved in lignin biosynthesis pathway, such as hydroxycinnamoyl CoA: shikimate hydroxycinnamoyl transferase (HCT), cinnamoyl CoA reductase (CCR), and cinnamyl alcohol dehydrogenase (CAD), through “terminator-less” constructs to reduce lignin in transgenic rice. Real-time qPCR analyses of the selected T1 transgenic rice plants indicated 36–86% transcript reduction in HCT lines, 75–94% in CCR lines, and 10–85% in CAD lines. Of the nine down-regulated lines (three lines from each genes) subjected to lignin analysis, seven showed significant reduction in total lignin content (HCT-4, HCT-7, CAD-1, CAD-7, CCR-3, CCR-7, and CCR-12) with lignin reduction ranging from 4.6 to 10.8%. The results from this study indicated that truncated gene fragments lacking transcription termination sequence can be used for down-regulation of lignin genes in rice, and the rice straw from these transgenic lines could be useful as feedstock for cellulosic biofuel.
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Linh D. Nguyen,Vibha Srivastava,Jamie L. Underwood,Soumen Nandy,M. Aydın Akbudak 한국식물생명공학회 2014 Plant biotechnology reports Vol.8 No.6
Efficient methods for DNA excision are neededfor removing selectable marker genes from transgenicplants. The present work evaluated the enhanced FLPrecombinase, FLPe, for excising FLP recombination target(FRT)-flanked marker genes, and generating marker-freerice lines. Previously, the transient FLPe activity was foundto be at least threefold higher on the transgene locuscompared to that of FLPwt, the wild-type FLP recombinase. In this study, transgenic plants expressing FLPe werecross-pollinated with the plants harboring FRT site toanalyze marker excision in F1 plants, and the transmissionof marker-free locus to F2 progeny. The FLPe activity,expressed by the strong promoter (maize ubiquitin-1 gene),efficiently excised FRT-flanked marker gene in rice plants. However, marker excision in F2 progeny was tightly linkedwith the presence of FLPe gene, suggesting insufficientrecombination in the gametophyte. The maize ubiquitin-1promoter is reportedly active in gametophytic tissue andeffective in meiotic transmission of the marker-free locusgenerated by Cre–lox recombination. Therefore, theobserved lack of meiotic transmission in this study ispossibly due to the limited efficiency of FLPe recombinase. While the reason for the FLPe inefficiency in the gametophyteis not clear, this work highlights the constraints ofFLPe recombinase in generating stable marker-free plantlines through cross-pollination or gene induction methods.
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