Jianxun Lin

Boston University, Boston, Massachusetts, United States

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Publications (6)19.43 Total impact

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    ABSTRACT: The optipore approach to nanopore-based sequencing enables direct optical readout of many DNA sequences in parallel using an array of nanopores. Fluorescent beacons hybridized to template strands of DNA threaded through each nanopore are sequentially excited via a 4-color total internal reflection (TIR) system. Here we present fabrication of a chip designed specifically for integration into the TIR-optipore illumination scheme. In the large freestanding silicon nitride membrane we have fabricated an array of small (sub 5 nm) nanopores with uniform geometry and optically resolvable (1.5 μm) spacing. We demonstrate that this chip design can be used for parallel readout from the 4-color optipore sequencer.
    No preview · Conference Paper · May 2013
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    ABSTRACT: Following epical completion of human genome project, further effort in sequencing arena was geared towards a steep reduction in cost to enable personalized genomic-based diagnostics. Nanopore devices provide an appealing platform for DNA sequencing at the single-molecule level, which enables direct recognition of minute amount of genome fragments without the hassle of enzymatic amplification. Classic electrical readout in nanopore sensing is still limited by its throughput as well as further downscaling of operation cost. In this demonstration, we presented a nanopore-array sensor equipped with optical readout modality on a custom prism-based TIRF microscope to allow simultaneous detection of DNA molecules passing through different nanopores under an electrical field. The oligonucleotide sequence to be deciphered was first expanded into a longer strand through a biochemical conversion procedure and each nucleotide unit was encoded into a short oligonucleotide strand with unique sequence. Molecular beacons (MBs) of different types were synthesized, each of which contains a short recognition sequence for each nucleotide base and a fluorescent reporter, and then hybridized with the converted oligonucleotide strand to form a duplex. The double-stranded oligonucleotides were driven through the nanopore electrically and the MB moieties were stripped off in a sequential manner and the identity of each original nucleotide base was then detected. Optical readout of fluorescence signal arising from each nucleotide can be performed at different nanopores simultaneously on a single high frame-rate CCD without need to fabricate sophisticated electrical circuitry to address each individual nanopore and holds a great promise for parallelized detection in DNA sequencing. We demonstrate that optical signals from different nanopores can be simultaneously read and identities of nucleotides from different oligonucleotide strands can be revealed in a parallel fashion.
    Full-text · Conference Paper · May 2013
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    Jianxun Lin · Marc Fabian · Nahum Sonenberg · Amit Meller
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    ABSTRACT: The ubiquitous and abundant cytoplasmic poly(A) binding protein (PABP) is a highly conserved multifunctional protein, many copies of which bind to the poly(A) tail of eukaryotic mRNAs to promote translation initiation. The N-terminus of PABP is responsible for the high binding specificity and affinity to poly(A), whereas the C-terminus is known to stimulate PABP multimerization on poly(A). Here, we use single-molecule nanopore force spectroscopy to directly measure interactions between poly(A) and PABPs. Both electrical and biochemical results show that the C-C domain interaction between two consecutive PABPs promotes cooperative binding. Up to now, investigators have not been able to probe the detailed polarity configuration (i.e., the internal arrangement of two PABPs on a poly(A) streak in which the C-termini face toward or away from each other). Our nanopore force spectroscopy system is able to distinguish the cooperative binding conformation from the noncooperative one. The ∼50% cooperative binding conformation of wild-type PABPs indicates that the C-C domain interaction doubles the cooperative binding probability. Moreover, the longer dissociation time of a cooperatively bound poly(A)/PABP complex as compared with a noncooperatively bound one indicates that the cooperative mode is the most stable conformation for PABPs binding onto the poly(A). However, ∼50% of the poly(A)/PABP complexes exhibit a noncooperative binding conformation, which is in line with previous studies showing that the PABP C-terminal domain also interacts with additional protein cofactors.
    Full-text · Article · Mar 2012 · Biophysical Journal
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    Jianxun Lin · Anatoly Kolomeisky · Amit Meller
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    ABSTRACT: Helix-coil transition kinetics of polyadenylic acid [poly(A)] inside a small protein channel is investigated for the first time, at the single molecule level. The confinement of a RNA molecule inside the channel slows its kinetics by nearly 3 orders of magnitude as compared to bulk measurements of free poly(A). These findings are related to the interaction energy of the RNA structure with the interior of the pore, explained by a simple two-state model. These results shed light on the way intermolecular interactions alter nucleic acid kinetics.
    Full-text · Article · Apr 2010 · Physical Review Letters
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    Jianxun Lin · Anatoly Kolomeisky · Amit Meller

    Full-text · Article · Jan 2010 · Biophysical Journal
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    Jianxun Lin · Ralf Bundschuh · Amit Meller

    Preview · Article · Feb 2009 · Biophysical Journal