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Research paper : Development of novel chemical reagents for reliable genetic analyses (Y. Komatsu et al.)−4−Synthesiology - English edition Vol.4 No.1 (2011) investigating the high reactivity of the ssN-linker along with the work of product realization and the basic research turned out to be fruitful. Soon, we found that the structure of the linker alters the chemical property of the neighboring primary amine increasing the reaction efficiency with the target molecule[4]. Since this structure was a new discovery not included in the patent for the first-generation product, we applied for a new patent, and this new structure was called the “second-generation amino-linker”. Although several compounds belong to the second-generation type due to the common structure, we selected the highly stable “ssH-linker” as the next linker after various investigations (Fig. 3). Unlike the ssN-linker, the ssH-linker does not contain the hydrophobic group in the molecule, but it is possible to very quickly remove the hydrophobic group that protected the amino group under moderate conditions. This chemical property of the ssH-linker enabled the high-throughput purification easily by reverse-phase chromatography, and it was also confirmed that the conjugation efficiency of this linker with the target molecule was superior to the conventional amino-linkers (Table 1, Fig. 5). Moreover, the ssH-linker was chemically more stable than the ssN-linker, and had the advantage of the synthesis cost being almost the same as the conventional reagent (Table1).Due to such superiority, we re-proposed the ssH-linker to the DNA synthesis company and the DNA chip manufacturing company, and requested the restart of the interrupted projects. Since it became possible to keep the unit price significantly low for the ssH-linker, the chemical synthesis company accepted it smoothly, while time was needed for getting approval from the DNA chip company that had been collecting data for the DNA chip using the ssN-linker. Later, the technological superiority of the ssH-linker was acknowledged by the two companies. We concluded a licensing agreement for the domestic DNA synthesis using the ssH-linker in 2006 with the DNA synthesis company. Also, in 2007 and 2008, the DNA chips using the ssH-modified probes were commercially produced.The ssH-linker has higher reactivity to active esters compared to conventional amino-linkers, and its modified DNAs and RNAs show high-purity by high-throughput purification. In addition, the low cost of the reagent promoted the use of the linker in the DNA synthesis companies, and it is now on market worldwide by overseas chemical companies. The license of the lower priced ssH-linker meant the reduction of royalties for us, but we decided to license it so that as many companies and research institutes use our product as possible.4.1.6 Development after licensingThe conventional amino linker has been used throughout the world for a long time, and it has been already built into the current genetic analysis systems. Therefore, it was not easy to replace the conventional linker by the new type having a different structure. This meant that after the license, we faced the difficulty of “selling a product or getting people to buy it.” To steadily increase the use of our linker, we continued application researches to propose new usages other than genetic analysis, as well as scientific demonstrations through publication of papers. As a result, we were able to propose an alternative usage of this reagent in the oligonucleotide therapeutic field[5], and the demand for linkers is recently AromaticgroupConventionalHighest activity:ssN-linkerOne of second-generation linkersFirst-generationDNAL2L1H2NssH-linkerNHOOOOH2NH2NIntroduction of aromatic groupImprovement of reactivity and purificationImportance of main chain structureImprovement of stabilityTechnologytransferUniversity, company, etc.Technologytransfer 1(DNA chip etc.)Amino linker④User①Synthesis ofreagentPractical useUserBiological evaluationSynthesis, purificationDevelopment and evaluation at AIST (small scale)Reagent synthesiscompanyGenetic analysiscompanydcbaDNA synthesiscompanyBusinessTechnologytransfer 2②DNA synthesis③Genetic analysissystemTechnologytransferJoint researchTechnologytransferJoint researchBusinessBusinessReagent synthesiscompanyDNA synthesiscompanyGenetic analysiscompanyFig. 3 Structures of the amino linkers bonded to the DNA and the flow of developmentFig. 4 Process from the linker reagent to the utilization of analysis systemBasic development and the construction and evaluation of the model system were conducted at AIST, and the efficacy of the performance was confirmed (b). Later, the technology was transferred to the private sector (c), evaluation of biological experiments were done at the genetic analysis company (c), and the route for utilizing the amino-linker reagent in bioresearch was established. Businesses are conducted for the reagent and products treated with the reagent.

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