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  • 95
    New England Biolabs restriction sites bamhi
    Sperm transfected with E6/E7 plasmid penetrate oocytes. a . Scheme of the recombinant plasmid pIRES2-AcGFP1-E6E7. E6/E7 genes have been amplified (1032 bp) from plasmid p1321 HPV-16 E6/E7 by PCR and subcloned to plasmid pIRES2-AcGFP1 between <t>SalI</t> and <t>BamHI</t> restriction sites. b . PCR for HPV E6/E7 genes from transfected sperm. Lane M: DNA marker (100 bp); 1: sperm transfected with recombinant E6/E7 plasmid; 2: negative control (no template); 3: sperm transfected only with Lipofectamine 2000; 4: positive control (pIRES2-AcGFP1-E6E7 plasmid). c . Mean number of human sperm penetrated per hamster oocyte in control and sperm transfected with HPV-16 E6/E7 plasmid. d . Hamster oocytes penetrated by control sperm and sperm transfected with HPV16 E6/E7 plasmid in bright field (BF, upper panel) and fluorescence (FL, lower panel) using SYBR green DNA stain.
    Restriction Sites Bamhi, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 149 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher bamhi restriction site
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
    Bamhi Restriction Site, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 97 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    TaKaRa bamhi restriction sites
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
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    92
    GenScript bamhi restriction sites
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
    Bamhi Restriction Sites, supplied by GenScript, used in various techniques. Bioz Stars score: 92/100, based on 142 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    System Biosciences Inc bamhi restriction sites
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
    Bamhi Restriction Sites, supplied by System Biosciences Inc, used in various techniques. Bioz Stars score: 92/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    ATUM bamh1 restriction sites
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
    Bamh1 Restriction Sites, supplied by ATUM, used in various techniques. Bioz Stars score: 93/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    GenScript xbai bamhi restriction sites
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
    Xbai Bamhi Restriction Sites, supplied by GenScript, used in various techniques. Bioz Stars score: 85/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    TaKaRa restriction enzyme sites bamhi ecori
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
    Restriction Enzyme Sites Bamhi Ecori, supplied by TaKaRa, used in various techniques. Bioz Stars score: 93/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    New England Biolabs ndei bamhi restriction sites
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
    Ndei Bamhi Restriction Sites, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 24 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Shanghai GenePharma bamh i restriction sites
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
    Bamh I Restriction Sites, supplied by Shanghai GenePharma, used in various techniques. Bioz Stars score: 93/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Genewiz bamhi restriction sites
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
    Bamhi Restriction Sites, supplied by Genewiz, used in various techniques. Bioz Stars score: 94/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    ProteoGenix restriction site bamhi
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
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    TaKaRa restriction sites bamhi
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
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    Promega bamh i restriction sites
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
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    Stratagene bamhi restriction site
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
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    Millipore bamhi restriction site
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
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    Eurofins bamhi restriction sites
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
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    Millipore restriction sites bamhi
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
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    Bio Basic Canada bamhi restriction sites
    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus <t>RNase</t> III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the <t>BamHI</t> recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.
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    Image Search Results


    Sperm transfected with E6/E7 plasmid penetrate oocytes. a . Scheme of the recombinant plasmid pIRES2-AcGFP1-E6E7. E6/E7 genes have been amplified (1032 bp) from plasmid p1321 HPV-16 E6/E7 by PCR and subcloned to plasmid pIRES2-AcGFP1 between SalI and BamHI restriction sites. b . PCR for HPV E6/E7 genes from transfected sperm. Lane M: DNA marker (100 bp); 1: sperm transfected with recombinant E6/E7 plasmid; 2: negative control (no template); 3: sperm transfected only with Lipofectamine 2000; 4: positive control (pIRES2-AcGFP1-E6E7 plasmid). c . Mean number of human sperm penetrated per hamster oocyte in control and sperm transfected with HPV-16 E6/E7 plasmid. d . Hamster oocytes penetrated by control sperm and sperm transfected with HPV16 E6/E7 plasmid in bright field (BF, upper panel) and fluorescence (FL, lower panel) using SYBR green DNA stain.

    Journal: PLoS ONE

    Article Title: Mechanism of Human Papillomavirus Binding to Human Spermatozoa and Fertilizing Ability of Infected Spermatozoa

    doi: 10.1371/journal.pone.0015036

    Figure Lengend Snippet: Sperm transfected with E6/E7 plasmid penetrate oocytes. a . Scheme of the recombinant plasmid pIRES2-AcGFP1-E6E7. E6/E7 genes have been amplified (1032 bp) from plasmid p1321 HPV-16 E6/E7 by PCR and subcloned to plasmid pIRES2-AcGFP1 between SalI and BamHI restriction sites. b . PCR for HPV E6/E7 genes from transfected sperm. Lane M: DNA marker (100 bp); 1: sperm transfected with recombinant E6/E7 plasmid; 2: negative control (no template); 3: sperm transfected only with Lipofectamine 2000; 4: positive control (pIRES2-AcGFP1-E6E7 plasmid). c . Mean number of human sperm penetrated per hamster oocyte in control and sperm transfected with HPV-16 E6/E7 plasmid. d . Hamster oocytes penetrated by control sperm and sperm transfected with HPV16 E6/E7 plasmid in bright field (BF, upper panel) and fluorescence (FL, lower panel) using SYBR green DNA stain.

    Article Snippet: The PCR mixture consisted of 5 µL Expand high fidelity buffer with MgCl2 , 1 µL of PCR grade nucleotide mix (10×), 20 pmol of each primer with SalI and BamHI restriction sites (New England Biolabs, Ipswich, MA) including forward: 5′-GTGCGCGTCGACGTGATGCACCAAAAGAGAACTG-3′ and reverse: 5′-GTGCGCGGATCCGTGTGGTTTCTGAGAACAGATG-3′ , 0,75 µL Expand high fidelity enzyme mix (Expand High Fidelity PCR System dNTPack, Roche Applied Science, Mannheim, Germany) and sterile H2 O in a final volume of 50 μL.

    Techniques: Transfection, Plasmid Preparation, Recombinant, Amplification, Polymerase Chain Reaction, Marker, Negative Control, Positive Control, Fluorescence, SYBR Green Assay, Staining

    Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus RNase III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the BamHI recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.

    Journal: Scientific Reports

    Article Title: Digital Imprinting of RNA Recognition and Processing on a Self-Assembled Nucleic Acid Matrix

    doi: 10.1038/srep02550

    Figure Lengend Snippet: Digital imprinting of dsRNA recognition and processing on a ds[RNA-DNA] matrix. Shown in (a) and (d) is the sequence of the 39-bp ds[RNA-DNA] (12-bp of dsRNA, 27-bp of dsDNA), containing an A. aeolicus RNase III processing site positioned in the middle of the dsRNA segment (|dsRNA 0 〉. The dsRNA sequence corresponds to the duplex stem in the A. aeolicus 23S rRNA precursor 38 . The red rectangles indicate sites of enzyme contact 38 , and the scissile phosphodiesters are indicated by the red arrows. (a) Input 1 (red): RNase III cleavage of the target site would create a surface-attached, 23-bp ds[RNA-DNA] with a 2-nt, 3′-overhang terminus (calculated length ~7.1 nm). Input 2 (magenta), uses the RNase III E110A mutant, which retains dsRNA-binding ability 31 39 . This input does not alter the height, and is essentially identical to the control experiment that omits RNase III (Input 3: green) (see also Table 1 ). (b) The H OUT /H IN value indicates that Output 1 corresponds to a ds[RNA-DNA] that is processed by RNase III (red dashed line), whereas Outputs 2 and 3 correspond to an unaltered dsDNA-dsRNA segment (black dashed line). (c) schematically depicts the output matrices [1–3], in which [1] consists of a monolayer of duplexes comprising the unaltered |dsDNA 0 〉 segment and a cleaved |dsRNA 0 〉 segment. The structures in (d) highlight the BamHI recognition sequence (enclosed in blue rectangles). BamHI action would create a surface-bound, 4-bp product possessing a 4-nt 3′-overhang (calculated length ~1.4 nm). In (e), three distinct ‘imprints’ are generated when BamHI is included (Inputs 1+–3+). For Input 1+, the ds[RNA-DNA] is processed at the BamHI site (blue dashed line), when RNase III also is present. For Input 3+, BamHI displays only limited catalytic action in the absence of RNase III, while for Input 2+, BamHI is inhibited in the presence of the E110A RNase III mutant (note error bars for Outputs 2+ and 3+). (f) schematically depicts the output matrices [1–3+], in which [1+] consists of a monolayer of duplexes comprising the cleaved |dsDNA 0 〉 segment, and [3+] consists of a mixed monolayer of cleaved |dsDNA 0 〉 segments and intact duplexes. Data are means, including the standard deviations.

    Article Snippet: Preparation of RNA-DNA chimera sequences A thiolated, 39-nucleotide RNA-DNA chimera sequence containing an RNase III cleavage site within the 27-nt RNA segment and a BamHI restriction site within the 12-nt DNA segment (see Seq1 below), as well as the complementary, non-thiolated [RNA-DNA] sequence (see Seq2 below) were purchased from ThermoFisher Scientific in HPLC-purified form.

    Techniques: Sequencing, Mutagenesis, Binding Assay, Generated

    Density-dependent steric regulation of imprinting a ds[RNA-DNA] matrix. (a) The final heights (H OUT ) of six separate Inputs are dependent upon the initial height (H IN ) of the ds[RNA-DNA] matrix. Input 1 (with RNase III). Input 2 (with E110A). Input 3 (controls, either lacking RNase III or with RNase III without the catalytic cofactor, Mg 2+ ). Input 1+ (with RNase III and BamHI). Input 2+ (with E110A and BamHI). Input 3+ (with BamHI alone). All dashed lines in (a) relate the data points to a linear regression. The data for Output 1 show that RNase III can process the dsRNA segment regardless of ds[RNA-DNA] density, which is related to the initial height (see schematic representation on top). Outputs 2 and 3 are consistent with an unaltered ds[RNA-DNA] chimera (represented by the solid diagonal line: H OUT = H IN ). BamHI gains full access to its site in combination with RNase III (Output 1+) as H OUT 1+ ≪ H OUT 1 , while it is essentially completely blocked in combination with the E110A mutant (Output 2+) as H OUT 2+ ~ H IN . BamHI restriction enzyme efficiency acting alone (Input 3+) must be lower than that of RNase III alone (Input 1), as the height of an matrix consisting of ds[RNA-DNA] molecules cleaved by BamHI would be lower than the height of an matrix cleaved by RNase III, and, in contrast, H OUT 3+ > H OUT 1 for relatively dense matrices (H IN > 10 nm). Data are means, and include standard deviations. (b) Schematic depiction of the effect of different inputs on a highly dense ds[RNA-DNA] matrix, including a steric hindrance-based model that shows how the ‘imprint’ (Output n+) is a step (i.e. digital) function of Input n+ (n = 1,3), as shown in (a).

    Journal: Scientific Reports

    Article Title: Digital Imprinting of RNA Recognition and Processing on a Self-Assembled Nucleic Acid Matrix

    doi: 10.1038/srep02550

    Figure Lengend Snippet: Density-dependent steric regulation of imprinting a ds[RNA-DNA] matrix. (a) The final heights (H OUT ) of six separate Inputs are dependent upon the initial height (H IN ) of the ds[RNA-DNA] matrix. Input 1 (with RNase III). Input 2 (with E110A). Input 3 (controls, either lacking RNase III or with RNase III without the catalytic cofactor, Mg 2+ ). Input 1+ (with RNase III and BamHI). Input 2+ (with E110A and BamHI). Input 3+ (with BamHI alone). All dashed lines in (a) relate the data points to a linear regression. The data for Output 1 show that RNase III can process the dsRNA segment regardless of ds[RNA-DNA] density, which is related to the initial height (see schematic representation on top). Outputs 2 and 3 are consistent with an unaltered ds[RNA-DNA] chimera (represented by the solid diagonal line: H OUT = H IN ). BamHI gains full access to its site in combination with RNase III (Output 1+) as H OUT 1+ ≪ H OUT 1 , while it is essentially completely blocked in combination with the E110A mutant (Output 2+) as H OUT 2+ ~ H IN . BamHI restriction enzyme efficiency acting alone (Input 3+) must be lower than that of RNase III alone (Input 1), as the height of an matrix consisting of ds[RNA-DNA] molecules cleaved by BamHI would be lower than the height of an matrix cleaved by RNase III, and, in contrast, H OUT 3+ > H OUT 1 for relatively dense matrices (H IN > 10 nm). Data are means, and include standard deviations. (b) Schematic depiction of the effect of different inputs on a highly dense ds[RNA-DNA] matrix, including a steric hindrance-based model that shows how the ‘imprint’ (Output n+) is a step (i.e. digital) function of Input n+ (n = 1,3), as shown in (a).

    Article Snippet: Preparation of RNA-DNA chimera sequences A thiolated, 39-nucleotide RNA-DNA chimera sequence containing an RNase III cleavage site within the 27-nt RNA segment and a BamHI restriction site within the 12-nt DNA segment (see Seq1 below), as well as the complementary, non-thiolated [RNA-DNA] sequence (see Seq2 below) were purchased from ThermoFisher Scientific in HPLC-purified form.

    Techniques: Mutagenesis