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  • 99
    Thermo Fisher verso cdna synthesis kit
    Differential expressions of genes modulating the mechanism of resistance to A. alternata in naturally infected mango fruits. (A) Effect of HWB on alternaria black spot (ABS) symptom development on mango cvs. Palmer, Kent, Tommy Atkins, Keitt, Lily and Shelly. (B) ABS symptom development on naturally infected fruits cv. Keitt following HWB treatment. (C) qRT-PCR differential expression profiling of genes Syn121 , glutardoxin , IT1K2 and AOS of cv. Shelly. Fruit peel tissues were sampled at four different time points after HWB treatment. <t>RNA</t> was extracted and served as a template for <t>cDNA</t> followed by qRT-PCR analysis of the genes of interest. Proportional increases in relative expression values were normalized against the samples of untreated mango fruits at 0 h. Expression data are means of two replicates. ABS-covered area was evaluated after 4 weeks of storage at 12°C. Average values followed by different letters differ significantly at P
    Verso Cdna Synthesis Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 6408 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Millipore xbai
    Linear view of consensus optical maps with <t>DNA</t> sequence-based maps. Solid black arrows represent the locations of missing cuts in the consensus optical maps. The <t>XbaI</t> and NheI consensus optical maps extend to the left of the origin of the DNA sequence-based map because both optical maps had a missing cut at that position.
    Xbai, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 715 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher amplitaq gold dna polymerase
    Marker TSC0033440 (G/C polymorphism). Exo I- and SAP-treated PCR products, genotyped without ddNTPs, re-using <t>AmpliTaq</t> Gold, from the PCR step on three pre-genotyped <t>DNA</t> samples. Reverse informative primer (5622 Da) was used for the primer extension with 25 µM dNTP depleted of dGTP. The expected extension products were 5936 Da (pA extension) and 6538 Da (pApCpA extension) for the C and G alleles, respectively. ( A ) Homozygous C/C sample for TSC0033440. ( B ) Heterozygous G/C sample for TSC0033440. ( C ) Homozygous G/G sample for TSC0033440.
    Amplitaq Gold Dna Polymerase, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 13081 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher superscript iii reverse transcriptase
    mRNAs stabilized in lsm mutants are 5′ capped. Immunoprecipitation of total RNAs with monoclonal anti-cap antibodies followed by <t>qRT-PCR</t> analysis of selected AtLSM1 (At4g11280, At5g62520, At5g12020, At4g32020), AtLSM8 (At4g11280, At5g62520) and AtXRN4 (At4g11280, At5g62520, At4g32020) substrates. Histograms represent the level of immunoprecipitated transcripts, where the value for Col-0 was arbitrarily set to 100. Values with standard deviations (±SD) were obtained from <t>three</t> independent experiments; * P
    Superscript Iii Reverse Transcriptase, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 90655 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher phusion dna polymerase
    Proportion of correct reads for the three genetic systems (simple: a single allele per individual, squares ; medium: two alleles, circles ; and complex: multiple alleles, triangles ) using standard PCR conditions ( open ) and modified PCR conditions to reduce chimera formation ( gray ). The size of the shape is indicative of the number of reads (see legend). All enzymes yielded at least 50% correct reads in the simplest system, mitochondrial <t>DNA</t> (Test 1; open squares). Some enzymes only worked for a given set of conditions (cycling conditions/genetic system). A group of enzymes consisting of <t>Phusion,</t> Gold and FastStart yielded a high proportion of correct reads cosistently accross all conditions. Others, such as Roche Taq, HotStar and Biotaq, yielded a low percent of correct reads for the more complex systems (MHC class I and MHC class II). Abbreviations as defined in Table 1 .
    Phusion Dna Polymerase, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 11361 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    TaKaRa la taq dna polymerase
    Proportion of correct reads for the three genetic systems (simple: a single allele per individual, squares ; medium: two alleles, circles ; and complex: multiple alleles, triangles ) using standard PCR conditions ( open ) and modified PCR conditions to reduce chimera formation ( gray ). The size of the shape is indicative of the number of reads (see legend). All enzymes yielded at least 50% correct reads in the simplest system, mitochondrial <t>DNA</t> (Test 1; open squares). Some enzymes only worked for a given set of conditions (cycling conditions/genetic system). A group of enzymes consisting of <t>Phusion,</t> Gold and FastStart yielded a high proportion of correct reads cosistently accross all conditions. Others, such as Roche Taq, HotStar and Biotaq, yielded a low percent of correct reads for the more complex systems (MHC class I and MHC class II). Abbreviations as defined in Table 1 .
    La Taq Dna Polymerase, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 5238 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher high capacity cdna reverse transcription kit
    Proportion of correct reads for the three genetic systems (simple: a single allele per individual, squares ; medium: two alleles, circles ; and complex: multiple alleles, triangles ) using standard PCR conditions ( open ) and modified PCR conditions to reduce chimera formation ( gray ). The size of the shape is indicative of the number of reads (see legend). All enzymes yielded at least 50% correct reads in the simplest system, mitochondrial <t>DNA</t> (Test 1; open squares). Some enzymes only worked for a given set of conditions (cycling conditions/genetic system). A group of enzymes consisting of <t>Phusion,</t> Gold and FastStart yielded a high proportion of correct reads cosistently accross all conditions. Others, such as Roche Taq, HotStar and Biotaq, yielded a low percent of correct reads for the more complex systems (MHC class I and MHC class II). Abbreviations as defined in Table 1 .
    High Capacity Cdna Reverse Transcription Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 117891 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher sybr green pcr master mix
    Real-time <t>PCR</t> amplification and dissociation (melt) curve plots. B. anthracis Melt-MAMA <t>SYBR®</t> Green assay targeting the A.Br.004 genetic clade. (A C) The amplification of two alleles are illustrated for haploid template ( Bacillus anthracis ) possessing an ‘A’ polymorphic SNP-state or ‘G’ state. Each amplification plot represents a single PCR reaction containing a reverse “common” primer and two allele-specific MAMA primers. The AS-MAMA primers anneal to the same template target and then compete for extension across the SNP position. The polymerase-mediated extension rate of the 3′match AS-MAMA primer (perfect primer-template complex) exceeds that of the 3′mismatched MAMA primer (mismatched primer-template complex), thus the perfect match primer-template complex outcompetes the mismatched primer-template complex and dominates the PCR amplification. (B D) Plots of the temperature-dissociation (melt) curve of the final PCR products for the two allele templates are shown next to their respective amplification plots (green arrows). Allele-specific PCR products are easily differentiated through temperature-dissociation (melt) curve analysis, which is conferred by the GC-clamp engineered on one of the AS-MAMA primer.
    Sybr Green Pcr Master Mix, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 101787 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher phusion hot start ii high fidelity dna polymerase
    Real-time <t>PCR</t> amplification and dissociation (melt) curve plots. B. anthracis Melt-MAMA <t>SYBR®</t> Green assay targeting the A.Br.004 genetic clade. (A C) The amplification of two alleles are illustrated for haploid template ( Bacillus anthracis ) possessing an ‘A’ polymorphic SNP-state or ‘G’ state. Each amplification plot represents a single PCR reaction containing a reverse “common” primer and two allele-specific MAMA primers. The AS-MAMA primers anneal to the same template target and then compete for extension across the SNP position. The polymerase-mediated extension rate of the 3′match AS-MAMA primer (perfect primer-template complex) exceeds that of the 3′mismatched MAMA primer (mismatched primer-template complex), thus the perfect match primer-template complex outcompetes the mismatched primer-template complex and dominates the PCR amplification. (B D) Plots of the temperature-dissociation (melt) curve of the final PCR products for the two allele templates are shown next to their respective amplification plots (green arrows). Allele-specific PCR products are easily differentiated through temperature-dissociation (melt) curve analysis, which is conferred by the GC-clamp engineered on one of the AS-MAMA primer.
    Phusion Hot Start Ii High Fidelity Dna Polymerase, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 2108 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    dntp  (TaKaRa)
    99
    TaKaRa dntp
    Real-time <t>PCR</t> amplification and dissociation (melt) curve plots. B. anthracis Melt-MAMA <t>SYBR®</t> Green assay targeting the A.Br.004 genetic clade. (A C) The amplification of two alleles are illustrated for haploid template ( Bacillus anthracis ) possessing an ‘A’ polymorphic SNP-state or ‘G’ state. Each amplification plot represents a single PCR reaction containing a reverse “common” primer and two allele-specific MAMA primers. The AS-MAMA primers anneal to the same template target and then compete for extension across the SNP position. The polymerase-mediated extension rate of the 3′match AS-MAMA primer (perfect primer-template complex) exceeds that of the 3′mismatched MAMA primer (mismatched primer-template complex), thus the perfect match primer-template complex outcompetes the mismatched primer-template complex and dominates the PCR amplification. (B D) Plots of the temperature-dissociation (melt) curve of the final PCR products for the two allele templates are shown next to their respective amplification plots (green arrows). Allele-specific PCR products are easily differentiated through temperature-dissociation (melt) curve analysis, which is conferred by the GC-clamp engineered on one of the AS-MAMA primer.
    Dntp, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 15826 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    taq  (TaKaRa)
    98
    TaKaRa taq
    Real-time <t>PCR</t> amplification and dissociation (melt) curve plots. B. anthracis Melt-MAMA <t>SYBR®</t> Green assay targeting the A.Br.004 genetic clade. (A C) The amplification of two alleles are illustrated for haploid template ( Bacillus anthracis ) possessing an ‘A’ polymorphic SNP-state or ‘G’ state. Each amplification plot represents a single PCR reaction containing a reverse “common” primer and two allele-specific MAMA primers. The AS-MAMA primers anneal to the same template target and then compete for extension across the SNP position. The polymerase-mediated extension rate of the 3′match AS-MAMA primer (perfect primer-template complex) exceeds that of the 3′mismatched MAMA primer (mismatched primer-template complex), thus the perfect match primer-template complex outcompetes the mismatched primer-template complex and dominates the PCR amplification. (B D) Plots of the temperature-dissociation (melt) curve of the final PCR products for the two allele templates are shown next to their respective amplification plots (green arrows). Allele-specific PCR products are easily differentiated through temperature-dissociation (melt) curve analysis, which is conferred by the GC-clamp engineered on one of the AS-MAMA primer.
    Taq, supplied by TaKaRa, used in various techniques. Bioz Stars score: 98/100, based on 732 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher opti mem
    Real-time <t>PCR</t> amplification and dissociation (melt) curve plots. B. anthracis Melt-MAMA <t>SYBR®</t> Green assay targeting the A.Br.004 genetic clade. (A C) The amplification of two alleles are illustrated for haploid template ( Bacillus anthracis ) possessing an ‘A’ polymorphic SNP-state or ‘G’ state. Each amplification plot represents a single PCR reaction containing a reverse “common” primer and two allele-specific MAMA primers. The AS-MAMA primers anneal to the same template target and then compete for extension across the SNP position. The polymerase-mediated extension rate of the 3′match AS-MAMA primer (perfect primer-template complex) exceeds that of the 3′mismatched MAMA primer (mismatched primer-template complex), thus the perfect match primer-template complex outcompetes the mismatched primer-template complex and dominates the PCR amplification. (B D) Plots of the temperature-dissociation (melt) curve of the final PCR products for the two allele templates are shown next to their respective amplification plots (green arrows). Allele-specific PCR products are easily differentiated through temperature-dissociation (melt) curve analysis, which is conferred by the GC-clamp engineered on one of the AS-MAMA primer.
    Opti Mem, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 56618 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher pcr buffer
    Silver-stained DGGE pattern of <t>PCR-amplified</t> 16S rDNA from the total <t>DNA</t> extracted from the deteriorated EBPR reactor. Electrophoresis was done for 2 to 6 h with a 1-h interval to optimize the running time. Up to 11 bands were clearly observed in the original gel. The six most dominant bands, which were further isolated and sequenced, are indicated next to the 3-h lane.
    Pcr Buffer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 28828 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher phusion high fidelity pcr master mix
    Silver-stained DGGE pattern of <t>PCR-amplified</t> 16S rDNA from the total <t>DNA</t> extracted from the deteriorated EBPR reactor. Electrophoresis was done for 2 to 6 h with a 1-h interval to optimize the running time. Up to 11 bands were clearly observed in the original gel. The six most dominant bands, which were further isolated and sequenced, are indicated next to the 3-h lane.
    Phusion High Fidelity Pcr Master Mix, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1884 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher fast sybr green master mix
    Silver-stained DGGE pattern of <t>PCR-amplified</t> 16S rDNA from the total <t>DNA</t> extracted from the deteriorated EBPR reactor. Electrophoresis was done for 2 to 6 h with a 1-h interval to optimize the running time. Up to 11 bands were clearly observed in the original gel. The six most dominant bands, which were further isolated and sequenced, are indicated next to the 3-h lane.
    Fast Sybr Green Master Mix, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 23476 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    TaKaRa gc buffer
    Silver-stained DGGE pattern of <t>PCR-amplified</t> 16S rDNA from the total <t>DNA</t> extracted from the deteriorated EBPR reactor. Electrophoresis was done for 2 to 6 h with a 1-h interval to optimize the running time. Up to 11 bands were clearly observed in the original gel. The six most dominant bands, which were further isolated and sequenced, are indicated next to the 3-h lane.
    Gc Buffer, supplied by TaKaRa, used in various techniques. Bioz Stars score: 96/100, based on 1235 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Differential expressions of genes modulating the mechanism of resistance to A. alternata in naturally infected mango fruits. (A) Effect of HWB on alternaria black spot (ABS) symptom development on mango cvs. Palmer, Kent, Tommy Atkins, Keitt, Lily and Shelly. (B) ABS symptom development on naturally infected fruits cv. Keitt following HWB treatment. (C) qRT-PCR differential expression profiling of genes Syn121 , glutardoxin , IT1K2 and AOS of cv. Shelly. Fruit peel tissues were sampled at four different time points after HWB treatment. RNA was extracted and served as a template for cDNA followed by qRT-PCR analysis of the genes of interest. Proportional increases in relative expression values were normalized against the samples of untreated mango fruits at 0 h. Expression data are means of two replicates. ABS-covered area was evaluated after 4 weeks of storage at 12°C. Average values followed by different letters differ significantly at P

    Journal: BMC Genomics

    Article Title: De-novo assembly of mango fruit peel transcriptome reveals mechanisms of mango response to hot water treatment

    doi: 10.1186/1471-2164-15-957

    Figure Lengend Snippet: Differential expressions of genes modulating the mechanism of resistance to A. alternata in naturally infected mango fruits. (A) Effect of HWB on alternaria black spot (ABS) symptom development on mango cvs. Palmer, Kent, Tommy Atkins, Keitt, Lily and Shelly. (B) ABS symptom development on naturally infected fruits cv. Keitt following HWB treatment. (C) qRT-PCR differential expression profiling of genes Syn121 , glutardoxin , IT1K2 and AOS of cv. Shelly. Fruit peel tissues were sampled at four different time points after HWB treatment. RNA was extracted and served as a template for cDNA followed by qRT-PCR analysis of the genes of interest. Proportional increases in relative expression values were normalized against the samples of untreated mango fruits at 0 h. Expression data are means of two replicates. ABS-covered area was evaluated after 4 weeks of storage at 12°C. Average values followed by different letters differ significantly at P

    Article Snippet: Analysis by qRT-PCR Single-stranded cDNA was synthesized from 1 μg of total RNA by means of the Verso cDNA synthesis kit (Thermo Fisher Scientific, Waltham, MA, USA).

    Techniques: Infection, Quantitative RT-PCR, Expressing

    EK transcriptional activation affects CMT1 downstream expression. a Schematic representation of the CMT1 gene and the insertion of EK within exon 13. CMT1 exons are numbered and shown as blue boxes and introns as black lines. Primers used to amplify upstream and downstream CMT1 sequences from cDNA are marked by arrowheads and numbered (P1–P7). The red broken line indicates the splice acceptor site of exon 14 (based on 27). EK LTRs are marked by red arrowheads. b CMT1 upstream transcript is alternatively spliced showing intron 12 retention. cDNA was prepared from RNA extracted from flowers derived from WT Col, WT Ler, ddm1, kyp2 , cmt3, ago4 and rdr2 and subjected to PCR to amplify CMT1 RNA sequences using primer set 1 + 2 (ex10-F + ex11-R) or primer set 3 + 4 (ex11-F + ex13-R). Note that primer set 3 + 4 yielded two PCR fragments SP1 and SP2. Actin was used as a reference. PCRs were performed for 35 cycles except for actin (30 cycles). M indicates DNA molecular size markers. The predicted alternative spliced variants SP1 and SP2 are schematically shown below. c CMT1 downstream expression using primer set 5 + 6 (ex14-F + ex16-R). WT Ler gDNA was used as a reference for PCR product derived from RNA. Note the enhanced downstream CMT1 expression in kyp2 and cmt3 mutants. Actin was used as a reference. Ler genomic DNA (gDNA) was used to confirm amplification from RNA. PCRs were for 35 cycles except for actin (30 cycles). d Analysis of splicing out of the entire EK retroelement using primer set 6 + 7 (ex13-F + ex16-R) on both sides of the EK insertion site. Note lack of a PCR product in kyp2 and cmt3 mutant. Actin was used as a reference RNA. Col genomic DNA (gDNA) was used to confirm amplification from RNA. PCR was for 42 cycles except for actin (30 cycles)

    Journal: Epigenetics & Chromatin

    Article Title: CMT3 and SUVH4/KYP silence the exonic Evelknievel retroelement to allow for reconstitution of CMT1 mRNA

    doi: 10.1186/s13072-018-0240-y

    Figure Lengend Snippet: EK transcriptional activation affects CMT1 downstream expression. a Schematic representation of the CMT1 gene and the insertion of EK within exon 13. CMT1 exons are numbered and shown as blue boxes and introns as black lines. Primers used to amplify upstream and downstream CMT1 sequences from cDNA are marked by arrowheads and numbered (P1–P7). The red broken line indicates the splice acceptor site of exon 14 (based on 27). EK LTRs are marked by red arrowheads. b CMT1 upstream transcript is alternatively spliced showing intron 12 retention. cDNA was prepared from RNA extracted from flowers derived from WT Col, WT Ler, ddm1, kyp2 , cmt3, ago4 and rdr2 and subjected to PCR to amplify CMT1 RNA sequences using primer set 1 + 2 (ex10-F + ex11-R) or primer set 3 + 4 (ex11-F + ex13-R). Note that primer set 3 + 4 yielded two PCR fragments SP1 and SP2. Actin was used as a reference. PCRs were performed for 35 cycles except for actin (30 cycles). M indicates DNA molecular size markers. The predicted alternative spliced variants SP1 and SP2 are schematically shown below. c CMT1 downstream expression using primer set 5 + 6 (ex14-F + ex16-R). WT Ler gDNA was used as a reference for PCR product derived from RNA. Note the enhanced downstream CMT1 expression in kyp2 and cmt3 mutants. Actin was used as a reference. Ler genomic DNA (gDNA) was used to confirm amplification from RNA. PCRs were for 35 cycles except for actin (30 cycles). d Analysis of splicing out of the entire EK retroelement using primer set 6 + 7 (ex13-F + ex16-R) on both sides of the EK insertion site. Note lack of a PCR product in kyp2 and cmt3 mutant. Actin was used as a reference RNA. Col genomic DNA (gDNA) was used to confirm amplification from RNA. PCR was for 42 cycles except for actin (30 cycles)

    Article Snippet: One microgram of total RNA was used for cDNA production using the Verso cDNA Kit (Thermo Scientific) according to the manufacturer’s protocol.

    Techniques: Activation Assay, Expressing, Derivative Assay, Polymerase Chain Reaction, Amplification, Mutagenesis

    Expression analysis of mecA in JE2 and mutant strains. RNA was isolated from the exponentially grown (0.5–1.2 OD 600nm ) cultures treated with or without oxicillin (Ox) 4 μg/ml for 60 min in MHB. cDNA was prepared from RNA samples and used as template with appropriate primer sets, and relative expression levels were calculated as mentioned in the Materials and Methods section. Each bar shows the mean and standard deviation of values obtained from three replicates. The effect of mutation and oxacillin on expression of pbp2 and mecA was analyzed by performing multiple pairwise comparisons for pbp2 (uppercase letters) and mecA (lowercase letters) using ANOVA followed by Tukey's post-hoc test, and p -values

    Journal: Frontiers in Microbiology

    Article Title: RelQ Mediates the Expression of β-Lactam Resistance in Methicillin-Resistant Staphylococcus aureus

    doi: 10.3389/fmicb.2019.00339

    Figure Lengend Snippet: Expression analysis of mecA in JE2 and mutant strains. RNA was isolated from the exponentially grown (0.5–1.2 OD 600nm ) cultures treated with or without oxicillin (Ox) 4 μg/ml for 60 min in MHB. cDNA was prepared from RNA samples and used as template with appropriate primer sets, and relative expression levels were calculated as mentioned in the Materials and Methods section. Each bar shows the mean and standard deviation of values obtained from three replicates. The effect of mutation and oxacillin on expression of pbp2 and mecA was analyzed by performing multiple pairwise comparisons for pbp2 (uppercase letters) and mecA (lowercase letters) using ANOVA followed by Tukey's post-hoc test, and p -values

    Article Snippet: For relative quantification, 2 μg total RNA was reverse transcribed using a cDNA Synthesis Kit (Fermentas, USA), and 1 μl of 10x diluted cDNA was used as template in 20 μl reaction volume.

    Techniques: Expressing, Mutagenesis, Isolation, Standard Deviation

    Effect of relQ expression on mecA transcription in parent and mutant strains. Transcript level was monitored by RT-PCR. cDNA was prepared by reverse-transcription of the RNA samples isolated from exponentially grown cultures treated with or without oxicillin (Ox) 4 μg/ml for 60 min in MHB. The relative expression levels were calculated using the 2 - Δ Δ C T method. Each bar shows the mean and standard deviation of values obtained from three replicates. The effect of relQ expression on mecA transcription level (lowercase letters) in different strains was analyzed by performing multiple pairwise comparisons using ANOVA followed by Tukey's post-hoc test, and p -values

    Journal: Frontiers in Microbiology

    Article Title: RelQ Mediates the Expression of β-Lactam Resistance in Methicillin-Resistant Staphylococcus aureus

    doi: 10.3389/fmicb.2019.00339

    Figure Lengend Snippet: Effect of relQ expression on mecA transcription in parent and mutant strains. Transcript level was monitored by RT-PCR. cDNA was prepared by reverse-transcription of the RNA samples isolated from exponentially grown cultures treated with or without oxicillin (Ox) 4 μg/ml for 60 min in MHB. The relative expression levels were calculated using the 2 - Δ Δ C T method. Each bar shows the mean and standard deviation of values obtained from three replicates. The effect of relQ expression on mecA transcription level (lowercase letters) in different strains was analyzed by performing multiple pairwise comparisons using ANOVA followed by Tukey's post-hoc test, and p -values

    Article Snippet: For relative quantification, 2 μg total RNA was reverse transcribed using a cDNA Synthesis Kit (Fermentas, USA), and 1 μl of 10x diluted cDNA was used as template in 20 μl reaction volume.

    Techniques: Expressing, Mutagenesis, Reverse Transcription Polymerase Chain Reaction, Isolation, Standard Deviation

    Linear view of consensus optical maps with DNA sequence-based maps. Solid black arrows represent the locations of missing cuts in the consensus optical maps. The XbaI and NheI consensus optical maps extend to the left of the origin of the DNA sequence-based map because both optical maps had a missing cut at that position.

    Journal: Applied and Environmental Microbiology

    Article Title: Whole-Genome Shotgun Optical Mapping of Rhodospirillum rubrum

    doi: 10.1128/AEM.71.9.5511-5522.2005

    Figure Lengend Snippet: Linear view of consensus optical maps with DNA sequence-based maps. Solid black arrows represent the locations of missing cuts in the consensus optical maps. The XbaI and NheI consensus optical maps extend to the left of the origin of the DNA sequence-based map because both optical maps had a missing cut at that position.

    Article Snippet: Surface properties were assayed by digesting lambda DASH II bacteriophage DNA with 40 units of XbaI, HindIII, and NheI diluted in 200 μl of digestion buffer with 0.2% Triton X-100 (Sigma, St. Louis, MO) at 37°C to determine optimal digestion times, which ranged from 30 to 120 min.

    Techniques: Sequencing

    Whole-genome circular XbaI map of R. rubrum . The outermost circle (thick line) represents the consensus map created by Gentig from the single-molecule maps shown as arcs. The single-molecule maps were made from single DNA molecules digested with XbaI. The color ordering of the fragments is for contrast and is therefore random.

    Journal: Applied and Environmental Microbiology

    Article Title: Whole-Genome Shotgun Optical Mapping of Rhodospirillum rubrum

    doi: 10.1128/AEM.71.9.5511-5522.2005

    Figure Lengend Snippet: Whole-genome circular XbaI map of R. rubrum . The outermost circle (thick line) represents the consensus map created by Gentig from the single-molecule maps shown as arcs. The single-molecule maps were made from single DNA molecules digested with XbaI. The color ordering of the fragments is for contrast and is therefore random.

    Article Snippet: Surface properties were assayed by digesting lambda DASH II bacteriophage DNA with 40 units of XbaI, HindIII, and NheI diluted in 200 μl of digestion buffer with 0.2% Triton X-100 (Sigma, St. Louis, MO) at 37°C to determine optimal digestion times, which ranged from 30 to 120 min.

    Techniques:

    Comparisons of the XbaI, NheI, and HindIII optical maps to sequence data. (A, B, and C) Plots of optical map fragment sizes versus the DNA sequence-based map fragment sizes from the finished sequence for XbaI (A), NheI (B), and HindIII (C). The error bars represent the standard deviation of optical map fragment size about the mean. (D, E, F) Plots of the relative fragment sizing error [100 × (optical map fragment size - corresponding DNA sequence-based map fragment size)/corresponding DNA sequence-based map fragment size] versus DNA sequence-based map fragment size for XbaI ( D), NheI (E), and HindIII (F).

    Journal: Applied and Environmental Microbiology

    Article Title: Whole-Genome Shotgun Optical Mapping of Rhodospirillum rubrum

    doi: 10.1128/AEM.71.9.5511-5522.2005

    Figure Lengend Snippet: Comparisons of the XbaI, NheI, and HindIII optical maps to sequence data. (A, B, and C) Plots of optical map fragment sizes versus the DNA sequence-based map fragment sizes from the finished sequence for XbaI (A), NheI (B), and HindIII (C). The error bars represent the standard deviation of optical map fragment size about the mean. (D, E, F) Plots of the relative fragment sizing error [100 × (optical map fragment size - corresponding DNA sequence-based map fragment size)/corresponding DNA sequence-based map fragment size] versus DNA sequence-based map fragment size for XbaI ( D), NheI (E), and HindIII (F).

    Article Snippet: Surface properties were assayed by digesting lambda DASH II bacteriophage DNA with 40 units of XbaI, HindIII, and NheI diluted in 200 μl of digestion buffer with 0.2% Triton X-100 (Sigma, St. Louis, MO) at 37°C to determine optimal digestion times, which ranged from 30 to 120 min.

    Techniques: Sequencing, Standard Deviation

    Dendrogram based on Xba I PFGE types identified among 31 E. coli strains. The data were sorted by the UPGMA method. AMP-ampicillin, TET-tetracycline, SUT- trimethoprim/sulfamethoxazole, CLO-chloramphenicol, CFL-cephalothin, AMC-amoxicillin/clavulanic acid, LVX- levofloxacin, CIP-ciprofloxacin, GEN-gentamicin, CRO-ceftriaxone, CTX-cefotaxime. *Isolates that show intermediate susceptibility to cephalotin.

    Journal: Brazilian Journal of Microbiology

    Article Title: Antimicrobial resistance and genetic diversity of Escherichiacoli isolated from humans and foods

    doi: 10.1590/S1517-838246420130874

    Figure Lengend Snippet: Dendrogram based on Xba I PFGE types identified among 31 E. coli strains. The data were sorted by the UPGMA method. AMP-ampicillin, TET-tetracycline, SUT- trimethoprim/sulfamethoxazole, CLO-chloramphenicol, CFL-cephalothin, AMC-amoxicillin/clavulanic acid, LVX- levofloxacin, CIP-ciprofloxacin, GEN-gentamicin, CRO-ceftriaxone, CTX-cefotaxime. *Isolates that show intermediate susceptibility to cephalotin.

    Article Snippet: Data Analysis The Xba I (Sigma-Aldrich, USA) fingerprints were analyzed using GelCompar II software (Applied Maths, Kortrijk, Belgium).

    Techniques:

    Comparison of digestion pattern of eDNA and genomic DNA (by DNase I and restriction enzymes). Agarose gel (0.8%) showing Lane 1: Genomic DNA, Lane 2: Digestion of genomic DNA with DNase I, Lane 3: Digestion of genomic DNA with restriction enzymes ( Bam HI, Eco RI, Hin dIII, Pst I, Xba I, Xho I), Lane M: Molecular weight marker ( λ phage genome Eco RI /Hin dIII digest), Lane 4: Digestion of eDNA with restriction enzymes ( Bam HI, Eco RI, Hin dIII, Pst I, Xba I, Xho I), Lane 5: Digestion of eDNA with DNase I, Lane 6: Purified eDNA.

    Journal: The Scientific World Journal

    Article Title: Characterization of eDNA from the Clinical Strain Acinetobacter baumannii AIIMS 7 and Its Role in Biofilm Formation

    doi: 10.1100/2012/973436

    Figure Lengend Snippet: Comparison of digestion pattern of eDNA and genomic DNA (by DNase I and restriction enzymes). Agarose gel (0.8%) showing Lane 1: Genomic DNA, Lane 2: Digestion of genomic DNA with DNase I, Lane 3: Digestion of genomic DNA with restriction enzymes ( Bam HI, Eco RI, Hin dIII, Pst I, Xba I, Xho I), Lane M: Molecular weight marker ( λ phage genome Eco RI /Hin dIII digest), Lane 4: Digestion of eDNA with restriction enzymes ( Bam HI, Eco RI, Hin dIII, Pst I, Xba I, Xho I), Lane 5: Digestion of eDNA with DNase I, Lane 6: Purified eDNA.

    Article Snippet: Digestion of eDNA by DNase I and Restriction Enzymes To check the digestion pattern of eDNA and compare with genomic DNA, eDNA was digested with six different restriction enzymes Bam HI, Eco RI, Hin dIII, Pst I, Xba I, and Xho I (Sigma Aldrich).

    Techniques: Agarose Gel Electrophoresis, Molecular Weight, Marker, Purification

    Marker TSC0033440 (G/C polymorphism). Exo I- and SAP-treated PCR products, genotyped without ddNTPs, re-using AmpliTaq Gold, from the PCR step on three pre-genotyped DNA samples. Reverse informative primer (5622 Da) was used for the primer extension with 25 µM dNTP depleted of dGTP. The expected extension products were 5936 Da (pA extension) and 6538 Da (pApCpA extension) for the C and G alleles, respectively. ( A ) Homozygous C/C sample for TSC0033440. ( B ) Heterozygous G/C sample for TSC0033440. ( C ) Homozygous G/G sample for TSC0033440.

    Journal: Nucleic Acids Research

    Article Title: A novel MALDI-TOF based methodology for genotyping single nucleotide polymorphisms

    doi: 10.1093/nar/gng156

    Figure Lengend Snippet: Marker TSC0033440 (G/C polymorphism). Exo I- and SAP-treated PCR products, genotyped without ddNTPs, re-using AmpliTaq Gold, from the PCR step on three pre-genotyped DNA samples. Reverse informative primer (5622 Da) was used for the primer extension with 25 µM dNTP depleted of dGTP. The expected extension products were 5936 Da (pA extension) and 6538 Da (pApCpA extension) for the C and G alleles, respectively. ( A ) Homozygous C/C sample for TSC0033440. ( B ) Heterozygous G/C sample for TSC0033440. ( C ) Homozygous G/G sample for TSC0033440.

    Article Snippet: AmpliTaq Gold® DNA polymerase was purchased from Applied Biosystems (Foster City, CA).

    Techniques: Marker, Polymerase Chain Reaction

    mRNAs stabilized in lsm mutants are 5′ capped. Immunoprecipitation of total RNAs with monoclonal anti-cap antibodies followed by qRT-PCR analysis of selected AtLSM1 (At4g11280, At5g62520, At5g12020, At4g32020), AtLSM8 (At4g11280, At5g62520) and AtXRN4 (At4g11280, At5g62520, At4g32020) substrates. Histograms represent the level of immunoprecipitated transcripts, where the value for Col-0 was arbitrarily set to 100. Values with standard deviations (±SD) were obtained from three independent experiments; * P

    Journal: Nucleic Acids Research

    Article Title: Arabidopsis thaliana LSM proteins function in mRNA splicing and degradation

    doi: 10.1093/nar/gkt296

    Figure Lengend Snippet: mRNAs stabilized in lsm mutants are 5′ capped. Immunoprecipitation of total RNAs with monoclonal anti-cap antibodies followed by qRT-PCR analysis of selected AtLSM1 (At4g11280, At5g62520, At5g12020, At4g32020), AtLSM8 (At4g11280, At5g62520) and AtXRN4 (At4g11280, At5g62520, At4g32020) substrates. Histograms represent the level of immunoprecipitated transcripts, where the value for Col-0 was arbitrarily set to 100. Values with standard deviations (±SD) were obtained from three independent experiments; * P

    Article Snippet: RNA samples, 500 ng of the input fraction and 60 ng of the IP fraction, were reverse transcribed for 60 min at 55°C with random primers and SuperScript III Reverse Transcriptase (Invitrogen) and used for PCR amplification.

    Techniques: Immunoprecipitation, Quantitative RT-PCR

    RNA-seq data displayed in Artemis. Mapped RNA-seq data is displayed as a plot showing sequence depth for the forward (blue) and reverse strand (red). The S. bongori genome annotation is also shown. The graphs, from the top downwards, represent the result of sequencing ( i ) undepleted ss-cDNA ( ii ) depleted ss-cDNA ( iii ) depleted ss-cDNA with actD present in the reverse transcription reaction ( iv ) ds-cDNA and (v) ds-cDNA with actD present in the reverse transcription reaction.

    Journal: Nucleic Acids Research

    Article Title: A simple method for directional transcriptome sequencing using Illumina technology

    doi: 10.1093/nar/gkp811

    Figure Lengend Snippet: RNA-seq data displayed in Artemis. Mapped RNA-seq data is displayed as a plot showing sequence depth for the forward (blue) and reverse strand (red). The S. bongori genome annotation is also shown. The graphs, from the top downwards, represent the result of sequencing ( i ) undepleted ss-cDNA ( ii ) depleted ss-cDNA ( iii ) depleted ss-cDNA with actD present in the reverse transcription reaction ( iv ) ds-cDNA and (v) ds-cDNA with actD present in the reverse transcription reaction.

    Article Snippet: RNA was denatured at 70°C for 10 min in the presence of 3 µg µl−1 random hexamer primers, then cooled on ice for 5 min. cDNA was then synthesized through reverse transcription using SuperScript III (Invitrogen) at 42°C for 2 h. When specified, actinomycin D (actD) (Sigma) was added to this reaction at a concentration of 6 µg ml−1 .

    Techniques: RNA Sequencing Assay, Sequencing

    Proportion of correct reads for the three genetic systems (simple: a single allele per individual, squares ; medium: two alleles, circles ; and complex: multiple alleles, triangles ) using standard PCR conditions ( open ) and modified PCR conditions to reduce chimera formation ( gray ). The size of the shape is indicative of the number of reads (see legend). All enzymes yielded at least 50% correct reads in the simplest system, mitochondrial DNA (Test 1; open squares). Some enzymes only worked for a given set of conditions (cycling conditions/genetic system). A group of enzymes consisting of Phusion, Gold and FastStart yielded a high proportion of correct reads cosistently accross all conditions. Others, such as Roche Taq, HotStar and Biotaq, yielded a low percent of correct reads for the more complex systems (MHC class I and MHC class II). Abbreviations as defined in Table 1 .

    Journal: Scientific Reports

    Article Title: Effect of the enzyme and PCR conditions on the quality of high-throughput DNA sequencing results

    doi: 10.1038/srep08056

    Figure Lengend Snippet: Proportion of correct reads for the three genetic systems (simple: a single allele per individual, squares ; medium: two alleles, circles ; and complex: multiple alleles, triangles ) using standard PCR conditions ( open ) and modified PCR conditions to reduce chimera formation ( gray ). The size of the shape is indicative of the number of reads (see legend). All enzymes yielded at least 50% correct reads in the simplest system, mitochondrial DNA (Test 1; open squares). Some enzymes only worked for a given set of conditions (cycling conditions/genetic system). A group of enzymes consisting of Phusion, Gold and FastStart yielded a high proportion of correct reads cosistently accross all conditions. Others, such as Roche Taq, HotStar and Biotaq, yielded a low percent of correct reads for the more complex systems (MHC class I and MHC class II). Abbreviations as defined in Table 1 .

    Article Snippet: Taq polymerase A range of 13 high fidelity, regular, economy and premium Taq polymerase enzymes were selected: Biotaq® (Bioline, London, UK), FastStart® High Fidelity PCR System (Roche, Mannheim, Germany), AmpliTaq Gold® (Applied Biosystems, Warrington, UK), HotStarTaq® DNA Polymerase (Qiagen, Hilden, Gernamy), Phusion® High Fidelity DNA Polymerase (Finnzymes, Espoo, Finland), Taq DNA Polymerase (Roche, Maylan, France), i-MaxTM II DNA Polymerase (iNtRON Biotechnology, Seongnam, Korea), KAPA HiFi™ (Kapa Biosystems, Boston, USA), OneTaq ™ DNA Polymerase (New England Biolabs, Hitchin, UK), Vent® DNA Polymerase (New England Biolabs, Hitchin, UK), Deep Vent® DNA Polymerase (New England Biolabs, Hitchin, UK), Pwo® DNA Polymerase (Roche, Maylan, France) and Velocity DNA Polymerase (Bioline, London, UK) (abbreviated names in ).

    Techniques: Polymerase Chain Reaction, Modification

    Real-time PCR amplification and dissociation (melt) curve plots. B. anthracis Melt-MAMA SYBR® Green assay targeting the A.Br.004 genetic clade. (A C) The amplification of two alleles are illustrated for haploid template ( Bacillus anthracis ) possessing an ‘A’ polymorphic SNP-state or ‘G’ state. Each amplification plot represents a single PCR reaction containing a reverse “common” primer and two allele-specific MAMA primers. The AS-MAMA primers anneal to the same template target and then compete for extension across the SNP position. The polymerase-mediated extension rate of the 3′match AS-MAMA primer (perfect primer-template complex) exceeds that of the 3′mismatched MAMA primer (mismatched primer-template complex), thus the perfect match primer-template complex outcompetes the mismatched primer-template complex and dominates the PCR amplification. (B D) Plots of the temperature-dissociation (melt) curve of the final PCR products for the two allele templates are shown next to their respective amplification plots (green arrows). Allele-specific PCR products are easily differentiated through temperature-dissociation (melt) curve analysis, which is conferred by the GC-clamp engineered on one of the AS-MAMA primer.

    Journal: PLoS ONE

    Article Title: Melt Analysis of Mismatch Amplification Mutation Assays (Melt-MAMA): A Functional Study of a Cost-Effective SNP Genotyping Assay in Bacterial Models

    doi: 10.1371/journal.pone.0032866

    Figure Lengend Snippet: Real-time PCR amplification and dissociation (melt) curve plots. B. anthracis Melt-MAMA SYBR® Green assay targeting the A.Br.004 genetic clade. (A C) The amplification of two alleles are illustrated for haploid template ( Bacillus anthracis ) possessing an ‘A’ polymorphic SNP-state or ‘G’ state. Each amplification plot represents a single PCR reaction containing a reverse “common” primer and two allele-specific MAMA primers. The AS-MAMA primers anneal to the same template target and then compete for extension across the SNP position. The polymerase-mediated extension rate of the 3′match AS-MAMA primer (perfect primer-template complex) exceeds that of the 3′mismatched MAMA primer (mismatched primer-template complex), thus the perfect match primer-template complex outcompetes the mismatched primer-template complex and dominates the PCR amplification. (B D) Plots of the temperature-dissociation (melt) curve of the final PCR products for the two allele templates are shown next to their respective amplification plots (green arrows). Allele-specific PCR products are easily differentiated through temperature-dissociation (melt) curve analysis, which is conferred by the GC-clamp engineered on one of the AS-MAMA primer.

    Article Snippet: Each Melt-MAMA reaction mixture (7 µl or 10 µl) contained 1x SYBR Green PCR master mix (Applied Biosystems, Foster City, CA), two forward AS primers, a common reverse primer, and 1 µl of DNA template at ∼1ng/µl.

    Techniques: Real-time Polymerase Chain Reaction, Amplification, SYBR Green Assay, Polymerase Chain Reaction

    Silver-stained DGGE pattern of PCR-amplified 16S rDNA from the total DNA extracted from the deteriorated EBPR reactor. Electrophoresis was done for 2 to 6 h with a 1-h interval to optimize the running time. Up to 11 bands were clearly observed in the original gel. The six most dominant bands, which were further isolated and sequenced, are indicated next to the 3-h lane.

    Journal: Applied and Environmental Microbiology

    Article Title: Identification of a Novel Group of Bacteria in Sludge from a Deteriorated Biological Phosphorus Removal Reactor

    doi:

    Figure Lengend Snippet: Silver-stained DGGE pattern of PCR-amplified 16S rDNA from the total DNA extracted from the deteriorated EBPR reactor. Electrophoresis was done for 2 to 6 h with a 1-h interval to optimize the running time. Up to 11 bands were clearly observed in the original gel. The six most dominant bands, which were further isolated and sequenced, are indicated next to the 3-h lane.

    Article Snippet: This DNA preparation was used as a DNA template in a PCR performed with 1× PCR buffer (Gibco BRL, Gaithersburg, Md.) containing a 200 μM concentration of each of the deoxynucleoside triphosphates, 1.5 mM MgCl2 , a 0.1 μM concentration of each of the primers (Operon Technologies, Inc., Alameda, Calif.), and 2.5 U of Taq polymerase (Pharmacia Biotech Inc., Piscataway, N.J.) in a final volume of 100 μl.

    Techniques: Staining, Denaturing Gradient Gel Electrophoresis, Polymerase Chain Reaction, Amplification, Electrophoresis, Isolation