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梁子才, Hong-Yan Zhang, , Jianping Mao, Daixing Zhou, Yunhe Xu, HaÊkan Thonberg, Zicai Liang, * and Claes Wahlestedt
Nucleic Acids Research, 2003, Vol. 31, No.14 e72,-0001,():
-1年11月30日
A solution-based method, mRNA accessible site tagging (MAST), has been developed to map the accessible sites of any given mRNA in high throughput fashion. mRNA molecules were immobilized and hybridized to randomized oligonucleotide libraries. Oligonucleotides speci®cally hybridized to the mRNA were sequenced and found to be able to precisely define the accessible sites of the mRNA. A number of ways were used to validate the accessible sites defined by the MAST process. Mapping of rabbit b-globin mRNA demonstrates the efficacy and advantage of MAST over other technologies in identifying accessible sites. Antisense oligonucleotides designed according to the accessible site map of human RhoA and Renilla luciferase mRNA result in knockdown effects that are in good correlation with the degrees of accessibility. The MAST methodology can be applied to mRNA of any length using a universal protocol.
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【期刊论文】Chemical nanoprinting: a novel method for fabricating DNA microchips
梁子才, Anil Kumar and Zicai Liang*
Nucleic Acids Research, 2001, Vol. 29, No.2 e2,-0001,():
-1年11月30日
We have developed a novel cost-effective procedure, namely 'chemical nanoprinting', for oligonucleotide or cDNA chips manufacture. In this thermo-controlled process, the oligonucleotides, covalently attached to a highly loaded 'master-chip' through disulfide bonds, are chemically transferred to the acrylamide layer mounted on a 'print-chip'. It is demonstrated here that multiple identical print-chips can be produced from a single master-chip. This duplication process is a few hundreds of times faster than any existing methods and the speed of process and cost incurred are independent of the scale of the DNA chips.
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【期刊论文】Vector-based siRNA delivery strategies for high-throughput screening of novel target genes
梁子才, Meihong Chen, Quan Du, Hong-Yan Zhang, Claes Wahlestedt, and Zicai Liang*
Journal of RNAi and Gene Silencing (2005), 1(1), 5-11,-0001,():
-1年11月30日
Application of siRNA in high-throughput fashion is still in its early phase although the principle has been established for three years. In this review, we outline the different vector-based siRNA delivery platforms as well as resources that are becoming available for high-throughput applications, and some initial outcomes of vector siRNA high-throughput screening efforts using vector encoded siRNA. It is expected that further improvement of the siRNA technology and availability of the siRNA resources will help to materialize the potential of siRNA for functional genomics and drug target validation.
siRNA, RNAi, vectors, gene silencing, siRNA delivery
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【期刊论文】A universal plasmid library encoding all permutations of small interfering RNA
梁子才, Meihong Chen*†‡, Lishu Zhang*†‡, Hong-Yan Zhang‡§, Xiahui Xiong*†, Bo Wang*†, Quan Du§, Bing Lu*, Claes Wahlestedt§¶, and Zicai Liang*
PNAS February 15, 2005 vol. 102 no.7,-0001,():
-1年11月30日
Small interfering RNA (siRNA) is normally designed to silence preselected known genes. Such selections are inevitably prone to bias as a result of limited knowledge about the biological process, transcript identity, and functions. A library that contains all permutations of siRNA could avoid such problems. In this paper, it is shown that 5×107 siRNA-encoding plasmids can be constructed in a single tube by using vectors with two mutated RNA polymerase III promoters arranged in a convergent manner. Such a library was used to carry out genomewide screening of functional genes in a phenotype-driven manner. Multiple siRNAs that induce a significant increase of cell proliferation speed were identified.
random targeting, screening, vector, dual promoters
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【期刊论文】Locked nucleic acid (LNA) mediated improvements in siRNA stability and functionality
梁子才, Joacim Elmen*, Hakan Thonberg, Karl Ljungberg, Miriam Frieden, Majken Westergaard, Yunhe Xu, Britta Wahren, Zicai Liang, Henrik Ørum, Troels Koch and Claes Wahlestedt
Nucleic Acids Research, 2005, Vol. 33, No.1 439-447,-0001,():
-1年11月30日
Therapeutic application of the recently discovered small interfering RNA (siRNA) gene silencing phenomenon will be dependent on improvements in molecule bio-stability, specificity and delivery. To address these issues, we have systematically modified siRNA with the synthetic RNA-like high affinity nucleotide analogue, Locked Nucleic Acid (LNA). Here,weshowthat incorporation of LNAsubstantially enhances serum half-life of siRNA’s, which is a key requirement for therapeutic use. Moreover, we provide evidence that LNA is compatible with the intracellular siRNA machinery and can be used to reduce undesired, sequence-related off-target effects. LNA-modified siRNAs targeting the emerging disease SARS, show improved efficiency over unmodified siRNA on certain RNA motifs. The results from this study emphasize LNA’s promise in converting siRNA from a functional genomics technology to a therapeutic platform.
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