bac dna miniprep kit  (Zymo Research)


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    Name:
    ZR BAC DNA Miniprep Kit
    Description:
    The ZR BAC DNA Miniprep Kit efficiently isolates BAC plasmid DNA or other large plasmids e g PAC from E Coli using a procedure that is simple rapid user friendly and reliable It features a modified alkaline lysis protocol with color coded reagents that allow easy visualization of complete bacterial cell lysis and neutralization The innovative Zymo Spin IC XL columns are optimized for high yield endotoxin free plasmid DNA recovery BAC DNA purified using the ZR BAC DNA Miniprep Kit is ideal for sequencing PCR endonuclease digestion etc
    Catalog Number:
    d4048
    Price:
    None
    Applications:
    DNA Purification
    Size:
    25 units
    Category:
    Life Science Reagents and Media
    Buy from Supplier


    Structured Review

    Zymo Research bac dna miniprep kit
    ZR BAC DNA Miniprep Kit
    The ZR BAC DNA Miniprep Kit efficiently isolates BAC plasmid DNA or other large plasmids e g PAC from E Coli using a procedure that is simple rapid user friendly and reliable It features a modified alkaline lysis protocol with color coded reagents that allow easy visualization of complete bacterial cell lysis and neutralization The innovative Zymo Spin IC XL columns are optimized for high yield endotoxin free plasmid DNA recovery BAC DNA purified using the ZR BAC DNA Miniprep Kit is ideal for sequencing PCR endonuclease digestion etc
    https://www.bioz.com/result/bac dna miniprep kit/product/Zymo Research
    Average 95 stars, based on 12 article reviews
    Price from $9.99 to $1999.99
    bac dna miniprep kit - by Bioz Stars, 2020-02
    95/100 stars

    Images

    1) Product Images from "Bioinformatic Tools Identify Chromosome-Specific DNA Probes and Facilitate Risk Assessment by Detecting Aneusomies in Extra-embryonic Tissues"

    Article Title: Bioinformatic Tools Identify Chromosome-Specific DNA Probes and Facilitate Risk Assessment by Detecting Aneusomies in Extra-embryonic Tissues

    Journal: Current Genomics

    doi: 10.2174/138920212802510510

    Screen dump of the UCSC Genome Browser (GoldenPath) display showing the BAC ends mapped to the targeted region of chromosome 2: 92,269,115 bp to 92,335,426 bp. The alignment BAC clone end sequences with the draft sequence of the human genome places BAC RP11-100H17 (arrow) in a region comprised entirely of satellite DNA, while BAC RP11-469P16 (arrowhead) is predicted to contain a cluster of long interspersed repeated DNA sequences (LINEs)(circled).
    Figure Legend Snippet: Screen dump of the UCSC Genome Browser (GoldenPath) display showing the BAC ends mapped to the targeted region of chromosome 2: 92,269,115 bp to 92,335,426 bp. The alignment BAC clone end sequences with the draft sequence of the human genome places BAC RP11-100H17 (arrow) in a region comprised entirely of satellite DNA, while BAC RP11-469P16 (arrowhead) is predicted to contain a cluster of long interspersed repeated DNA sequences (LINEs)(circled).

    Techniques Used: BAC Assay, Sequencing

    Screen dump of the UCSC Genome Browser display showing the BAC ends mapped to the region chromosome 4: 49,093,534 bp to 49,183,369 bp. The alignment BAC clone end sequences with the draft sequence of the human genome places BAC RP11-360M1 (high-lighted) in a region comprised almost entirely of satellite DNA and completely free of interspersed repeated DNA sequences such as short interspersed repeats (SINEs) or LINEs.
    Figure Legend Snippet: Screen dump of the UCSC Genome Browser display showing the BAC ends mapped to the region chromosome 4: 49,093,534 bp to 49,183,369 bp. The alignment BAC clone end sequences with the draft sequence of the human genome places BAC RP11-360M1 (high-lighted) in a region comprised almost entirely of satellite DNA and completely free of interspersed repeated DNA sequences such as short interspersed repeats (SINEs) or LINEs.

    Techniques Used: BAC Assay, Sequencing

    In situ hybridization analysis of DNA probes prepared from BAC clones. The BAC clones RP11-294C12 and RP11-242E13 hybridized to metaphase spreads prepared from short term cultures of human lymphocytes showed specific hybridization to the target regions on the X (arrowhead) and Y (arrow) chromosome, respectively. A) Schematic representation of the FISH target regions on the X and Y chromosome. B) Hybridization of both probes to metaphase chromosomes. C) Hybridization signals in diploid interphase cell nuclei. (Bars = 10 µm).
    Figure Legend Snippet: In situ hybridization analysis of DNA probes prepared from BAC clones. The BAC clones RP11-294C12 and RP11-242E13 hybridized to metaphase spreads prepared from short term cultures of human lymphocytes showed specific hybridization to the target regions on the X (arrowhead) and Y (arrow) chromosome, respectively. A) Schematic representation of the FISH target regions on the X and Y chromosome. B) Hybridization of both probes to metaphase chromosomes. C) Hybridization signals in diploid interphase cell nuclei. (Bars = 10 µm).

    Techniques Used: In Situ Hybridization, BAC Assay, Clone Assay, Hybridization, Fluorescence In Situ Hybridization

    FISH performance of BAC-derived DNA probes targeting the centromeric heterochromatin of chromosome 2. A-B) A DNA probe prepared from BAC clone RP11-469P16 shows multiple signals in normal interphase cell nuclei ( A ) or on metaphase spreads ( B ). Arrows in B ) point at the target region on chromosome 2. C ) A DNA probe prepared from BAC clone RP11-100H17 binds exclusively to the chromosome 2-specific target region (arrows). (Bars = 10 µm).
    Figure Legend Snippet: FISH performance of BAC-derived DNA probes targeting the centromeric heterochromatin of chromosome 2. A-B) A DNA probe prepared from BAC clone RP11-469P16 shows multiple signals in normal interphase cell nuclei ( A ) or on metaphase spreads ( B ). Arrows in B ) point at the target region on chromosome 2. C ) A DNA probe prepared from BAC clone RP11-100H17 binds exclusively to the chromosome 2-specific target region (arrows). (Bars = 10 µm).

    Techniques Used: Fluorescence In Situ Hybridization, BAC Assay, Derivative Assay

    2) Product Images from "Bioinformatics Tools Allow Targeted Selection of Chromosome Enumeration Probes and Aneuploidy Detection"

    Article Title: Bioinformatics Tools Allow Targeted Selection of Chromosome Enumeration Probes and Aneuploidy Detection

    Journal: Journal of Histochemistry and Cytochemistry

    doi: 10.1369/0022155412470955

    Culture genotype of chromosome 10 in a PTC cell line. Chromosome numbers and signal counts of the chromosome 10-specific DNA probe prepared from BAC RP11-168p20 hybridized to cell line S48TK18A6 (scored by three independent scorers).
    Figure Legend Snippet: Culture genotype of chromosome 10 in a PTC cell line. Chromosome numbers and signal counts of the chromosome 10-specific DNA probe prepared from BAC RP11-168p20 hybridized to cell line S48TK18A6 (scored by three independent scorers).

    Techniques Used: BAC Assay

    In situ hybridization results for selected DNA probes. Red shaded areas on the idiograms in (A–B) and (F–G) indicate the hybridization target sites. (A) In situ hybridization using fluorescent probes for the highly repeated satellite III DNA segment (flanked by the PCR primers WYR2 and WYR4) shows bright signals on the long arm of the Y chromosome. (B) Combining biotinylated in situ hybridization probes for the Y chromosome-specific DNA segment (large signal) with probes for X chromosome-specific DNA flanked by the PCR primers WXR1 and WXR2 (small signal) allows chromosome identification based on the size of the signal. (C) A digoxigenin-labeled DNA probe prepared from BAC RP11-348g24 resulted in specific signals on the X chromosome in the presence of a significant amount of cross-hybridization to other chromosomes in metaphase spreads. High level of cross-hybridization was also evident in interphase nuclei. (D) In contrast to the result shown in (C), a probe prepared from BAC RP11-294c12 bound almost exclusively to the centromeric heterochromatin of the human X chromosome. (E) Combining differentially labeled DNA probes in a single hybridization experiment, the Y chromosomal target appears in green, whereas the X chromosomal centromeric repeat DNA is shown in red. (F) In situ hybridization of an autosome-targeting DNA probe prepared from BAC clone RP11-96f8 showed multiple signals and a high level of cross-hybridization in the interphase cell nuclei. (G) The biotinylated probe prepared from BAC RP11-168p20 exhibited strong signals on both homologues of chromosome 10. (H) A combination of three differently labeled probes was hybridized simultaneously. Chromosome 10 signals are shown in red; X chromosomal and Y chromosomal signals are shown in green and blue, respectively. (I–K) The same probe as in (G) (BAC RP11-168p20) was hybridized to the cell line S48TK18A6. Multiple signals indicate chromosomal abnormalities. (I) shows DAPI picture, (J) shows green avidin-FITC signal, and (K) shows the superposition of both. (L–N) A chromosome X probe (BAC clone RP11-294c12) labeled with biotin/avidin-FITC, a Cy-5 labeled chromosome Y probe (RP11-242e13), and a chromosome 10-specific probe prepared from BAC RP11-168p20 (labeled with Spectral Orange-dUTP) were hybridized onto a deparaffinized 12-week placenta tissue section. (L) The chromosome 10 probe (red signals) and bound chromosome X probe (green signals) in this female placental tissue section. (M–N) In our triple probe FISH experiment, the chromosome 10 probe is represented by orange signals, the chromosome X probe by green signals, and the chromosome Y probe by red signals in the male cell nuclei. (Size marker bars indicate 10 µm.)
    Figure Legend Snippet: In situ hybridization results for selected DNA probes. Red shaded areas on the idiograms in (A–B) and (F–G) indicate the hybridization target sites. (A) In situ hybridization using fluorescent probes for the highly repeated satellite III DNA segment (flanked by the PCR primers WYR2 and WYR4) shows bright signals on the long arm of the Y chromosome. (B) Combining biotinylated in situ hybridization probes for the Y chromosome-specific DNA segment (large signal) with probes for X chromosome-specific DNA flanked by the PCR primers WXR1 and WXR2 (small signal) allows chromosome identification based on the size of the signal. (C) A digoxigenin-labeled DNA probe prepared from BAC RP11-348g24 resulted in specific signals on the X chromosome in the presence of a significant amount of cross-hybridization to other chromosomes in metaphase spreads. High level of cross-hybridization was also evident in interphase nuclei. (D) In contrast to the result shown in (C), a probe prepared from BAC RP11-294c12 bound almost exclusively to the centromeric heterochromatin of the human X chromosome. (E) Combining differentially labeled DNA probes in a single hybridization experiment, the Y chromosomal target appears in green, whereas the X chromosomal centromeric repeat DNA is shown in red. (F) In situ hybridization of an autosome-targeting DNA probe prepared from BAC clone RP11-96f8 showed multiple signals and a high level of cross-hybridization in the interphase cell nuclei. (G) The biotinylated probe prepared from BAC RP11-168p20 exhibited strong signals on both homologues of chromosome 10. (H) A combination of three differently labeled probes was hybridized simultaneously. Chromosome 10 signals are shown in red; X chromosomal and Y chromosomal signals are shown in green and blue, respectively. (I–K) The same probe as in (G) (BAC RP11-168p20) was hybridized to the cell line S48TK18A6. Multiple signals indicate chromosomal abnormalities. (I) shows DAPI picture, (J) shows green avidin-FITC signal, and (K) shows the superposition of both. (L–N) A chromosome X probe (BAC clone RP11-294c12) labeled with biotin/avidin-FITC, a Cy-5 labeled chromosome Y probe (RP11-242e13), and a chromosome 10-specific probe prepared from BAC RP11-168p20 (labeled with Spectral Orange-dUTP) were hybridized onto a deparaffinized 12-week placenta tissue section. (L) The chromosome 10 probe (red signals) and bound chromosome X probe (green signals) in this female placental tissue section. (M–N) In our triple probe FISH experiment, the chromosome 10 probe is represented by orange signals, the chromosome X probe by green signals, and the chromosome Y probe by red signals in the male cell nuclei. (Size marker bars indicate 10 µm.)

    Techniques Used: In Situ Hybridization, Hybridization, Polymerase Chain Reaction, Labeling, BAC Assay, Avidin-Biotin Assay, Fluorescence In Situ Hybridization, Marker

    Related Articles

    Clone Assay:

    Article Title: Multiple organ gigantism caused by mutation in VmPPD gene in blackgram (Vigna mungo)
    Article Snippet: After screening, the BAC DNA of the clone VmR48p23 was extracted using ZR BAC DNA Miniprep Kit (Zymo Research). .. The extracted DNA was sheared into ~3 kb fragments, cloned into pUC118 vector, sequenced with BigDye Terminator (Thermo Fisher Scientific) and ABI3730 (Thermo Fisher Scientific), assembled by CLC Genomic Workbench (QIAGEN) and annotated manually.

    Article Title: Chromosome-Specific DNA Repeats: Rapid Identification in Silico and Validation Using Fluorescence in Situ Hybridization
    Article Snippet: .. Fluorescence in situ Hybridization Individual BAC clones were grown overnight in up to 10 mL of Luria broth (LB) medium containing 12.5 μg/mL chloramphenicol (Sigma, St. Louis, MO, USA), and the DNA was isolated using a ZR BAC DNA Miniprep Kit (Zymo Research, Irvine, CA, USA). .. For the preparation of DNA pools, clones were grown individually and pooled prior to DNA isolation.

    Article Title: Establishment of an Efficient and Flexible Genetic Manipulation Platform Based on a Fosmid Library for Rapid Generation of Recombinant Pseudorabies Virus
    Article Snippet: Therefore, 200 clones were randomly picked and cultured overnight in 5-ml LB liquid medium containing 12.5 μg/ml chloramphenicol and 50 μl of auto-induction solution (Epicentre). .. The fosmids were extracted using ZR BAC DNA Miniprep Kit (Zymo Research).

    Article Title: A human monoclonal antibody that specifically binds and inhibits the staphylococcal complement inhibitor protein SCIN
    Article Snippet: Expression of staphylococcal SCIN and CHIPS proteins in L. lactis Primers used for cloning are described in . .. Bacterial chromosomal DNA was isolated using the ZR BAC DNA Miniprep Kit (Zymo Reasearch Corporation, USA) following the manufacturer's protocol.

    Article Title: A PECTIN METHYLESTERASE gene at the maize Ga1 locus confers male function in unilateral cross-incompatibility
    Article Snippet: We developed a PCR-based method to screen clones harboring the pollen determinant. .. A 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA).

    Article Title: Wheat Ms2 encodes for an orphan protein that confers male sterility in grass species
    Article Snippet: In total, 706,176 BAC clones were arranged in 1,839 384-well plates, representing a 5.1-fold coverage of the wheat genome (∼17 Gb) ( ). .. In brief, a 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA).

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: Purification of Recombinant Xenopus CMG The open reading frames encoding the eleven X. laevis CMG subunits (CDC45, MCM2-7 hetero-hexamer, and GINS hetero-tetramer) were cloned into a single plasmid (pGC187) using the MultiBac system . .. The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research).

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: Codon-optimized Xenopus laevis cDNAs encoding all four GINS subunits (synthesized by IDT) were cloned into a single expression plasmid (pGC128) using the MultiBac system ( ). .. The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048).

    Amplification:

    Article Title: Colibactin assembly line enzymes use S-adenosylmethionine to build a cyclopropane ring
    Article Snippet: .. The mutant BACs were isolated from these colonies using ZR BAC DNA Miniprep Kit (Zymo research), and gene disruption was verified by PCR amplification of knockout regions. .. To generate an E. coli mutant strain expressing ClbA, ClbB, ClbC, and ClbN, E. coli DH10B were electroporated with mutant BACpks ΔclbDtoM ΔclbOtoQ and selected on LB plates containing 50 μg/mL apramycin and 50 μg/mL kanamycin.

    Article Title: A human monoclonal antibody that specifically binds and inhibits the staphylococcal complement inhibitor protein SCIN
    Article Snippet: DNA amplification was performed using Fusion Hot start High-Fidelity DNA polymerase according the instructions of the supplier (Thermoscientific). .. Bacterial chromosomal DNA was isolated using the ZR BAC DNA Miniprep Kit (Zymo Reasearch Corporation, USA) following the manufacturer's protocol.

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: MCM2-7 open reading frames were PCR amplified from X. laevis cDNA. .. The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research).

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048). .. Baculovirus encoding GINS was amplified in three stages (P1, P2, and P3) in Sf9 cells (Expression Systems Cat# 94–001S) and GINS expression levels were monitored via western blots.

    Article Title: Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA
    Article Snippet: Briefly, full-length viral cDNAs flanked by Not I sites were amplified by long RT-PCR using primers 5′Cstrain_T7_Not1 (which includes a T7 promotor for in vitro transcription, a Not I site and a region corresponding to the first 44 nt of the genome) and 3′CSFV_Not1 (that contains a Not I site and sequence complementary to the 3′-terminal 35 nt of the genome that are conserved among many CSFVs including the C-strain). .. BACs to be used as templates for long PCR or for screening by restriction enzyme digestion were purified from 4 ml overnight cultures of E. coli DH10B using the ZR BAC DNA Miniprep Kit (Zymo Research).

    Synthesized:

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: Codon-optimized GINS and CDC45 open reading frames were synthesized as gBlocks by IDT. .. The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research).

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: Codon-optimized Xenopus laevis cDNAs encoding all four GINS subunits (synthesized by IDT) were cloned into a single expression plasmid (pGC128) using the MultiBac system ( ). .. The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048).

    Construct:

    Article Title: Multiple organ gigantism caused by mutation in VmPPD gene in blackgram (Vigna mungo)
    Article Snippet: BAC library After we found the four gene loci were completely linked to the mog phenotype, we screened the BAC library constructed from genomic DNA of the blackgram cultivar ‘Phitsanulok 2’. .. After screening, the BAC DNA of the clone VmR48p23 was extracted using ZR BAC DNA Miniprep Kit (Zymo Research).

    Article Title: A PECTIN METHYLESTERASE gene at the maize Ga1 locus confers male function in unilateral cross-incompatibility
    Article Snippet: Construction and screening of the SDGa25 BAC library To construct a BAC library of SDGa25, high-molecular weight genomic DNA was extracted from leaf tissues, partially digested by the restriction enzyme Hin dIII, and ligated into the vector pIndigoBAC536-S , . .. A 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA).

    Article Title: Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA
    Article Snippet: Paragraph title: BAC constructs ... BACs to be used as templates for long PCR or for screening by restriction enzyme digestion were purified from 4 ml overnight cultures of E. coli DH10B using the ZR BAC DNA Miniprep Kit (Zymo Research).

    Infection:

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research). .. 500 mL of Tni (similar to HiFive) cell culture (Expression Systems) at a density of 2-3 million/mL was infected with 50 mL baculovirus (MOI > 10).

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048). .. 500mL of Tni cell culture (Expression Systems Cat# 94–002S) at a density of 2–3 was infected with 5–10 mL P3 baculovirus (MOI > 1).

    Expressing:

    Article Title: Colibactin assembly line enzymes use S-adenosylmethionine to build a cyclopropane ring
    Article Snippet: To generate an E. coli mutant strain expressing ClbA, ClbB, ClbC, ClbN, ClbO, ClbP, and ClbQ, E. coli DH10B were electroporated with mutant BACpks ΔclbDtoM and selected on LB plates containing 50 μg/mL apramycin and 50 μg/mL kanamycin. .. The mutant BACs were isolated from these colonies using ZR BAC DNA Miniprep Kit (Zymo research), and gene disruption was verified by PCR amplification of knockout regions.

    Article Title: A human monoclonal antibody that specifically binds and inhibits the staphylococcal complement inhibitor protein SCIN
    Article Snippet: Paragraph title: Expression of staphylococcal SCIN and CHIPS proteins in L. lactis ... Bacterial chromosomal DNA was isolated using the ZR BAC DNA Miniprep Kit (Zymo Reasearch Corporation, USA) following the manufacturer's protocol.

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research). .. The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research).

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: Paragraph title: Expressing, and purifying recombinant GINS ... The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048).

    Modification:

    Article Title: Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA
    Article Snippet: All BACs were modified and maintained in E. coli DH10B cells (Invitrogen) grown at 37°C in LB medium containing chloramphenicol (Cam, 15 μg/ml). .. BACs to be used as templates for long PCR or for screening by restriction enzyme digestion were purified from 4 ml overnight cultures of E. coli DH10B using the ZR BAC DNA Miniprep Kit (Zymo Research).

    Western Blot:

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048). .. Baculovirus encoding GINS was amplified in three stages (P1, P2, and P3) in Sf9 cells (Expression Systems Cat# 94–001S) and GINS expression levels were monitored via western blots.

    Transformation Assay:

    Article Title: A PECTIN METHYLESTERASE gene at the maize Ga1 locus confers male function in unilateral cross-incompatibility
    Article Snippet: The ligation products were transformed into ElectroMAX DH10B T1 phage-resistant cells (Invitrogene) and screened on LB medium with chloramphenicol (12.5 mg/L), 5-bromo-4-chloro-3-indolyl-β-d -galactopyranoside (80 mg/L) and isopropyl β-d -1-thiogalactoside (100 mg/L). .. A 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA).

    Article Title: Wheat Ms2 encodes for an orphan protein that confers male sterility in grass species
    Article Snippet: The ligation product was transformed into the ElectroMAX DH10B T1 phage-resistant cells (Invitrogene, Carlsbad, CA, USA) and screened on the LB medium with 12.5 mg l−1 chloramphenicol, 80 mg l−1 X-gal and 100 mg l−1 IPTG. .. In brief, a 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA).

    Derivative Assay:

    Article Title: Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA
    Article Snippet: BAC constructs The BAC construct, pBeloR26, was constructed using the long RT-PCR method as previously described [ ] using RNA derived from the “C-strain Riems”. .. BACs to be used as templates for long PCR or for screening by restriction enzyme digestion were purified from 4 ml overnight cultures of E. coli DH10B using the ZR BAC DNA Miniprep Kit (Zymo Research).

    Electroporation:

    Article Title: Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA
    Article Snippet: The electroporation of bacteria was performed in 0.1 cm cuvettes using 1 pulse at 1800 V, 25 μF and 200 Ω in a Gene Pulser Xcell (Bio-Rad). .. BACs to be used as templates for long PCR or for screening by restriction enzyme digestion were purified from 4 ml overnight cultures of E. coli DH10B using the ZR BAC DNA Miniprep Kit (Zymo Research).

    Transfection:

    Article Title: LANA oligomeric architecture is essential for KSHV nuclear body formation and viral genome maintenance during latency
    Article Snippet: BAC16 iSLK cells BAC16 and its derivatives were transfected into iSLK cells using the Effectene Transfection Reagent kit (QIAGEN, 301425). .. Briefly, BAC16 DNA was isolated from 2ml bacterial culture using a ZR BAC DNA Miniprep Kit (Zymo Research) and resuspended in 30 μl of RNase-free water. iSLK cells were seeded at 2x105 cells/well of a 6-well plate.

    Sequencing:

    Article Title: Multiple organ gigantism caused by mutation in VmPPD gene in blackgram (Vigna mungo)
    Article Snippet: After screening, the BAC DNA of the clone VmR48p23 was extracted using ZR BAC DNA Miniprep Kit (Zymo Research). .. The sequence of the assembled BAC clone was deposited as LC101731 in DNA Data Bank of Japan ( http://www.ddbj.nig.ac.jp/ ).

    Article Title: Establishment of an Efficient and Flexible Genetic Manipulation Platform Based on a Fosmid Library for Rapid Generation of Recombinant Pseudorabies Virus
    Article Snippet: Paragraph title: Fosmid Sequencing and Full-Length Genome Assembly ... The fosmids were extracted using ZR BAC DNA Miniprep Kit (Zymo Research).

    Article Title: A PECTIN METHYLESTERASE gene at the maize Ga1 locus confers male function in unilateral cross-incompatibility
    Article Snippet: A 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA). .. BAC DNA sequencing was performed using PacBio SMRT sequencing technology by the Nextomics Biosciences Company (Wuhan, China).

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: A sequence encoding the FLAG tag was added to the N-terminus of MCM3. .. The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research).

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: A sequence encoding a 10-aa linker, the LPETG tag for sortase labeling, and a His6 tag was added to the C-terminus of the Psf3 subunit (GGGGSGGGGS-LPETG-HHHHHH). .. The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048).

    Article Title: Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA
    Article Snippet: Briefly, full-length viral cDNAs flanked by Not I sites were amplified by long RT-PCR using primers 5′Cstrain_T7_Not1 (which includes a T7 promotor for in vitro transcription, a Not I site and a region corresponding to the first 44 nt of the genome) and 3′CSFV_Not1 (that contains a Not I site and sequence complementary to the 3′-terminal 35 nt of the genome that are conserved among many CSFVs including the C-strain). .. BACs to be used as templates for long PCR or for screening by restriction enzyme digestion were purified from 4 ml overnight cultures of E. coli DH10B using the ZR BAC DNA Miniprep Kit (Zymo Research).

    Ligation:

    Article Title: A human monoclonal antibody that specifically binds and inhibits the staphylococcal complement inhibitor protein SCIN
    Article Snippet: Bacterial chromosomal DNA was isolated using the ZR BAC DNA Miniprep Kit (Zymo Reasearch Corporation, USA) following the manufacturer's protocol. .. Cloning of the PCR-amplified scn and chp genes was performed by Not 1 and Bam H1 (New England Biolabs) cleavage followed by ligation to Not I/Bam HI cleft plasmid pNG4210.

    Article Title: A PECTIN METHYLESTERASE gene at the maize Ga1 locus confers male function in unilateral cross-incompatibility
    Article Snippet: The ligation products were transformed into ElectroMAX DH10B T1 phage-resistant cells (Invitrogene) and screened on LB medium with chloramphenicol (12.5 mg/L), 5-bromo-4-chloro-3-indolyl-β-d -galactopyranoside (80 mg/L) and isopropyl β-d -1-thiogalactoside (100 mg/L). .. A 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA).

    Article Title: Wheat Ms2 encodes for an orphan protein that confers male sterility in grass species
    Article Snippet: The ligation product was transformed into the ElectroMAX DH10B T1 phage-resistant cells (Invitrogene, Carlsbad, CA, USA) and screened on the LB medium with 12.5 mg l−1 chloramphenicol, 80 mg l−1 X-gal and 100 mg l−1 IPTG. .. In brief, a 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA).

    Protease Inhibitor:

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048). .. Cells harvested 48 hrs post-infection were pelleted at 500 × g for 15min and re-suspended in a final volume of 50mL in GINS Lysis Buffer (GLB) containing 20mM Tris-HCl pH8.0, 5% glycerol, 500mM NaCl, 20mM Imidazole, 1mM DTT, 1mM PMSF, EDTA-free cOmplete protease inhibitor cocktail (Roche Cat # 11873580001).

    Cell Culture:

    Article Title: Establishment of an Efficient and Flexible Genetic Manipulation Platform Based on a Fosmid Library for Rapid Generation of Recombinant Pseudorabies Virus
    Article Snippet: Therefore, 200 clones were randomly picked and cultured overnight in 5-ml LB liquid medium containing 12.5 μg/ml chloramphenicol and 50 μl of auto-induction solution (Epicentre). .. The fosmids were extracted using ZR BAC DNA Miniprep Kit (Zymo Research).

    Article Title: LANA oligomeric architecture is essential for KSHV nuclear body formation and viral genome maintenance during latency
    Article Snippet: Briefly, BAC16 DNA was isolated from 2ml bacterial culture using a ZR BAC DNA Miniprep Kit (Zymo Research) and resuspended in 30 μl of RNase-free water. iSLK cells were seeded at 2x105 cells/well of a 6-well plate. .. On following day, transfected cells were trypsinized and transferred into 10 cm dishes and cultured in DMEM supplemented with 10% FBS and 1% penicillin-streptomycin.

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research). .. 500 mL of Tni (similar to HiFive) cell culture (Expression Systems) at a density of 2-3 million/mL was infected with 50 mL baculovirus (MOI > 10).

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048). .. 500mL of Tni cell culture (Expression Systems Cat# 94–002S) at a density of 2–3 was infected with 5–10 mL P3 baculovirus (MOI > 1).

    Generated:

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research). .. CMG baculovirus was generated by transfecting pGC187 bacmid into Sf9 cells (Expression Systems) for 96 hr, and amplifying the virus in 2 rounds of 96 hr each.

    DNA Sequencing:

    Article Title: A PECTIN METHYLESTERASE gene at the maize Ga1 locus confers male function in unilateral cross-incompatibility
    Article Snippet: A 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA). .. BAC DNA sequencing was performed using PacBio SMRT sequencing technology by the Nextomics Biosciences Company (Wuhan, China).

    Polymerase Chain Reaction:

    Article Title: Colibactin assembly line enzymes use S-adenosylmethionine to build a cyclopropane ring
    Article Snippet: .. The mutant BACs were isolated from these colonies using ZR BAC DNA Miniprep Kit (Zymo research), and gene disruption was verified by PCR amplification of knockout regions. .. To generate an E. coli mutant strain expressing ClbA, ClbB, ClbC, and ClbN, E. coli DH10B were electroporated with mutant BACpks ΔclbDtoM ΔclbOtoQ and selected on LB plates containing 50 μg/mL apramycin and 50 μg/mL kanamycin.

    Article Title: A human monoclonal antibody that specifically binds and inhibits the staphylococcal complement inhibitor protein SCIN
    Article Snippet: Bacterial chromosomal DNA was isolated using the ZR BAC DNA Miniprep Kit (Zymo Reasearch Corporation, USA) following the manufacturer's protocol. .. Cloning of the PCR-amplified scn and chp genes was performed by Not 1 and Bam H1 (New England Biolabs) cleavage followed by ligation to Not I/Bam HI cleft plasmid pNG4210.

    Article Title: A PECTIN METHYLESTERASE gene at the maize Ga1 locus confers male function in unilateral cross-incompatibility
    Article Snippet: We developed a PCR-based method to screen clones harboring the pollen determinant. .. A 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA).

    Article Title: Wheat Ms2 encodes for an orphan protein that confers male sterility in grass species
    Article Snippet: We developed a PCR-based screening of the LM15RMs2 BAC library. .. In brief, a 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA).

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: MCM2-7 open reading frames were PCR amplified from X. laevis cDNA. .. The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research).

    Article Title: Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA
    Article Snippet: .. BACs to be used as templates for long PCR or for screening by restriction enzyme digestion were purified from 4 ml overnight cultures of E. coli DH10B using the ZR BAC DNA Miniprep Kit (Zymo Research). .. BACs required for direct genome sequencing were purified from 500 ml cultures using the Large-construct kit (Qiagen).

    Recombinant:

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: Paragraph title: Purification of Recombinant Xenopus CMG ... The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research).

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: Paragraph title: Expressing, and purifying recombinant GINS ... The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048).

    Molecular Weight:

    Article Title: Chromosome-Specific DNA Repeats: Rapid Identification in Silico and Validation Using Fluorescence in Situ Hybridization
    Article Snippet: Fluorescence in situ Hybridization Individual BAC clones were grown overnight in up to 10 mL of Luria broth (LB) medium containing 12.5 μg/mL chloramphenicol (Sigma, St. Louis, MO, USA), and the DNA was isolated using a ZR BAC DNA Miniprep Kit (Zymo Research, Irvine, CA, USA). .. The isolation of high molecular weight BAC DNAs was confirmed on 1% agarose gels and quantitated by spectrophotometry (Nanodrop 2000, Thermo Scientific, Wilmington, DE, USA).

    Article Title: Bioinformatics Tools Allow Targeted Selection of Chromosome Enumeration Probes and Aneuploidy Detection
    Article Snippet: DNA Probe Preparation The BAC DNAs were extracted from overnight cultures following an alkaline lysis protocol ( ) or using a ZR BAC DNA Miniprep Kit (Zymo Research; Irvine, CA). .. The isolation of high molecular weight BAC DNAs was confirmed on 1% agarose gels and quantitated by Hoechst fluorometry using a Hoefer TK 100 instrument (Hoefer; South San Francisco, CA).

    DNA Extraction:

    Article Title: Chromosome-Specific DNA Repeats: Rapid Identification in Silico and Validation Using Fluorescence in Situ Hybridization
    Article Snippet: Fluorescence in situ Hybridization Individual BAC clones were grown overnight in up to 10 mL of Luria broth (LB) medium containing 12.5 μg/mL chloramphenicol (Sigma, St. Louis, MO, USA), and the DNA was isolated using a ZR BAC DNA Miniprep Kit (Zymo Research, Irvine, CA, USA). .. For the preparation of DNA pools, clones were grown individually and pooled prior to DNA isolation.

    Fluorescence:

    Article Title: Chromosome-Specific DNA Repeats: Rapid Identification in Silico and Validation Using Fluorescence in Situ Hybridization
    Article Snippet: .. Fluorescence in situ Hybridization Individual BAC clones were grown overnight in up to 10 mL of Luria broth (LB) medium containing 12.5 μg/mL chloramphenicol (Sigma, St. Louis, MO, USA), and the DNA was isolated using a ZR BAC DNA Miniprep Kit (Zymo Research, Irvine, CA, USA). .. For the preparation of DNA pools, clones were grown individually and pooled prior to DNA isolation.

    Mutagenesis:

    Article Title: Colibactin assembly line enzymes use S-adenosylmethionine to build a cyclopropane ring
    Article Snippet: .. The mutant BACs were isolated from these colonies using ZR BAC DNA Miniprep Kit (Zymo research), and gene disruption was verified by PCR amplification of knockout regions. .. To generate an E. coli mutant strain expressing ClbA, ClbB, ClbC, and ClbN, E. coli DH10B were electroporated with mutant BACpks ΔclbDtoM ΔclbOtoQ and selected on LB plates containing 50 μg/mL apramycin and 50 μg/mL kanamycin.

    Isolation:

    Article Title: Chromosome-Specific DNA Repeats: Rapid Identification in Silico and Validation Using Fluorescence in Situ Hybridization
    Article Snippet: .. Fluorescence in situ Hybridization Individual BAC clones were grown overnight in up to 10 mL of Luria broth (LB) medium containing 12.5 μg/mL chloramphenicol (Sigma, St. Louis, MO, USA), and the DNA was isolated using a ZR BAC DNA Miniprep Kit (Zymo Research, Irvine, CA, USA). .. For the preparation of DNA pools, clones were grown individually and pooled prior to DNA isolation.

    Article Title: Colibactin assembly line enzymes use S-adenosylmethionine to build a cyclopropane ring
    Article Snippet: .. The mutant BACs were isolated from these colonies using ZR BAC DNA Miniprep Kit (Zymo research), and gene disruption was verified by PCR amplification of knockout regions. .. To generate an E. coli mutant strain expressing ClbA, ClbB, ClbC, and ClbN, E. coli DH10B were electroporated with mutant BACpks ΔclbDtoM ΔclbOtoQ and selected on LB plates containing 50 μg/mL apramycin and 50 μg/mL kanamycin.

    Article Title: A human monoclonal antibody that specifically binds and inhibits the staphylococcal complement inhibitor protein SCIN
    Article Snippet: .. Bacterial chromosomal DNA was isolated using the ZR BAC DNA Miniprep Kit (Zymo Reasearch Corporation, USA) following the manufacturer's protocol. ..

    Article Title: LANA oligomeric architecture is essential for KSHV nuclear body formation and viral genome maintenance during latency
    Article Snippet: .. Briefly, BAC16 DNA was isolated from 2ml bacterial culture using a ZR BAC DNA Miniprep Kit (Zymo Research) and resuspended in 30 μl of RNase-free water. iSLK cells were seeded at 2x105 cells/well of a 6-well plate. .. On the next day 15 μl of BAC DNA was mixed with transfection solution and added to the cells following by transfection kit’s according to manufacturer’s protocol.

    Article Title: Escape of X-linked miRNA genes from meiotic sex chromosome inactivation
    Article Snippet: .. BAC/fosmid DNA was isolated using the ZR BAC DNA miniprep Kit (ZymoResearch) according to the manufacturer's protocol. .. DNA concentration and quality were characterized by agarose gel electrophoresis and analysis on a BioRad Nanodrop.

    Article Title: Bioinformatics Tools Allow Targeted Selection of Chromosome Enumeration Probes and Aneuploidy Detection
    Article Snippet: DNA Probe Preparation The BAC DNAs were extracted from overnight cultures following an alkaline lysis protocol ( ) or using a ZR BAC DNA Miniprep Kit (Zymo Research; Irvine, CA). .. The isolation of high molecular weight BAC DNAs was confirmed on 1% agarose gels and quantitated by Hoechst fluorometry using a Hoefer TK 100 instrument (Hoefer; South San Francisco, CA).

    Labeling:

    Article Title: Chromosome-Specific DNA Repeats: Rapid Identification in Silico and Validation Using Fluorescence in Situ Hybridization
    Article Snippet: Fluorescence in situ Hybridization Individual BAC clones were grown overnight in up to 10 mL of Luria broth (LB) medium containing 12.5 μg/mL chloramphenicol (Sigma, St. Louis, MO, USA), and the DNA was isolated using a ZR BAC DNA Miniprep Kit (Zymo Research, Irvine, CA, USA). .. Probe DNAs were labeled with biotin-14-dCTP (part of the BioPrime kit, Invitrogen, La Jolla, CA, USA), digoxigenin-11-dUTP (Roche Diagnostics, Indianapolis, IN, USA) or Spectrum Green-dUTP (Abbott, Downers Grove, IL, USA) by random priming using a commercial kit (BioPrime Kit, Invitrogen) [ ].

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: A sequence encoding a 10-aa linker, the LPETG tag for sortase labeling, and a His6 tag was added to the C-terminus of the Psf3 subunit (GGGGSGGGGS-LPETG-HHHHHH). .. The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048).

    Article Title: Bioinformatics Tools Allow Targeted Selection of Chromosome Enumeration Probes and Aneuploidy Detection
    Article Snippet: DNA Probe Preparation The BAC DNAs were extracted from overnight cultures following an alkaline lysis protocol ( ) or using a ZR BAC DNA Miniprep Kit (Zymo Research; Irvine, CA). .. Probe DNAs were labeled with biotin-14-dCTP (Invitrogen; Carlsbad, CA), digoxigenin-11-dUTP (Roche Molecular Systems; Indianapolis, IN), Spectrum Orange-dUTP (Abbott; Abbott Park, IL), Spectrum Green-dUTP (Abbott), or Cy-5–dUTP (GE Healthcare; Piscataway Township, NJ) by random priming using a commercial kit (BioPrime Kit; Invitrogen) ( ; ).

    Purification:

    Article Title: Colibactin assembly line enzymes use S-adenosylmethionine to build a cyclopropane ring
    Article Snippet: To further generate a BACpks ΔclbDtoM ΔclbOtoQ mutant, the purified disruption cassette for clbOtoQ was electroporated into E. coli BW25113 harboring pKD46 and BACpks ΔclbDtoM . .. The mutant BACs were isolated from these colonies using ZR BAC DNA Miniprep Kit (Zymo research), and gene disruption was verified by PCR amplification of knockout regions.

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: .. The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research). .. CMG baculovirus was generated by transfecting pGC187 bacmid into Sf9 cells (Expression Systems) for 96 hr, and amplifying the virus in 2 rounds of 96 hr each.

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: .. The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048). .. Baculovirus encoding GINS was amplified in three stages (P1, P2, and P3) in Sf9 cells (Expression Systems Cat# 94–001S) and GINS expression levels were monitored via western blots.

    Article Title: Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA
    Article Snippet: .. BACs to be used as templates for long PCR or for screening by restriction enzyme digestion were purified from 4 ml overnight cultures of E. coli DH10B using the ZR BAC DNA Miniprep Kit (Zymo Research). .. BACs required for direct genome sequencing were purified from 500 ml cultures using the Large-construct kit (Qiagen).

    Reverse Transcription Polymerase Chain Reaction:

    Article Title: Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA
    Article Snippet: Briefly, full-length viral cDNAs flanked by Not I sites were amplified by long RT-PCR using primers 5′Cstrain_T7_Not1 (which includes a T7 promotor for in vitro transcription, a Not I site and a region corresponding to the first 44 nt of the genome) and 3′CSFV_Not1 (that contains a Not I site and sequence complementary to the 3′-terminal 35 nt of the genome that are conserved among many CSFVs including the C-strain). .. BACs to be used as templates for long PCR or for screening by restriction enzyme digestion were purified from 4 ml overnight cultures of E. coli DH10B using the ZR BAC DNA Miniprep Kit (Zymo Research).

    In Situ Hybridization:

    Article Title: Chromosome-Specific DNA Repeats: Rapid Identification in Silico and Validation Using Fluorescence in Situ Hybridization
    Article Snippet: .. Fluorescence in situ Hybridization Individual BAC clones were grown overnight in up to 10 mL of Luria broth (LB) medium containing 12.5 μg/mL chloramphenicol (Sigma, St. Louis, MO, USA), and the DNA was isolated using a ZR BAC DNA Miniprep Kit (Zymo Research, Irvine, CA, USA). .. For the preparation of DNA pools, clones were grown individually and pooled prior to DNA isolation.

    Plasmid Preparation:

    Article Title: Multiple organ gigantism caused by mutation in VmPPD gene in blackgram (Vigna mungo)
    Article Snippet: After screening, the BAC DNA of the clone VmR48p23 was extracted using ZR BAC DNA Miniprep Kit (Zymo Research). .. The extracted DNA was sheared into ~3 kb fragments, cloned into pUC118 vector, sequenced with BigDye Terminator (Thermo Fisher Scientific) and ABI3730 (Thermo Fisher Scientific), assembled by CLC Genomic Workbench (QIAGEN) and annotated manually.

    Article Title: Colibactin assembly line enzymes use S-adenosylmethionine to build a cyclopropane ring
    Article Snippet: To generate a BACpks ΔclbDtoM mutant, the purified disruption cassette for clbDtoM was electroporated into E. coli BW25113 harboring the λ Red recombinase expression plasmid pkD46 and BACpks . .. The mutant BACs were isolated from these colonies using ZR BAC DNA Miniprep Kit (Zymo research), and gene disruption was verified by PCR amplification of knockout regions.

    Article Title: A human monoclonal antibody that specifically binds and inhibits the staphylococcal complement inhibitor protein SCIN
    Article Snippet: Bacterial chromosomal DNA was isolated using the ZR BAC DNA Miniprep Kit (Zymo Reasearch Corporation, USA) following the manufacturer's protocol. .. Cloning of the PCR-amplified scn and chp genes was performed by Not 1 and Bam H1 (New England Biolabs) cleavage followed by ligation to Not I/Bam HI cleft plasmid pNG4210.

    Article Title: A PECTIN METHYLESTERASE gene at the maize Ga1 locus confers male function in unilateral cross-incompatibility
    Article Snippet: Construction and screening of the SDGa25 BAC library To construct a BAC library of SDGa25, high-molecular weight genomic DNA was extracted from leaf tissues, partially digested by the restriction enzyme Hin dIII, and ligated into the vector pIndigoBAC536-S , . .. A 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA).

    Article Title: Wheat Ms2 encodes for an orphan protein that confers male sterility in grass species
    Article Snippet: Briefly, high-molecular weight genomic DNA was extracted from leaf tissues, partially digested by the restriction enzyme Hind III, and ligated into the BAC vector pIndigoBAC536-S . .. In brief, a 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA).

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: Purification of Recombinant Xenopus CMG The open reading frames encoding the eleven X. laevis CMG subunits (CDC45, MCM2-7 hetero-hexamer, and GINS hetero-tetramer) were cloned into a single plasmid (pGC187) using the MultiBac system . .. The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research).

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: Codon-optimized Xenopus laevis cDNAs encoding all four GINS subunits (synthesized by IDT) were cloned into a single expression plasmid (pGC128) using the MultiBac system ( ). .. The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048).

    In Vitro:

    Article Title: Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA
    Article Snippet: Briefly, full-length viral cDNAs flanked by Not I sites were amplified by long RT-PCR using primers 5′Cstrain_T7_Not1 (which includes a T7 promotor for in vitro transcription, a Not I site and a region corresponding to the first 44 nt of the genome) and 3′CSFV_Not1 (that contains a Not I site and sequence complementary to the 3′-terminal 35 nt of the genome that are conserved among many CSFVs including the C-strain). .. BACs to be used as templates for long PCR or for screening by restriction enzyme digestion were purified from 4 ml overnight cultures of E. coli DH10B using the ZR BAC DNA Miniprep Kit (Zymo Research).

    Knock-Out:

    Article Title: Colibactin assembly line enzymes use S-adenosylmethionine to build a cyclopropane ring
    Article Snippet: .. The mutant BACs were isolated from these colonies using ZR BAC DNA Miniprep Kit (Zymo research), and gene disruption was verified by PCR amplification of knockout regions. .. To generate an E. coli mutant strain expressing ClbA, ClbB, ClbC, and ClbN, E. coli DH10B were electroporated with mutant BACpks ΔclbDtoM ΔclbOtoQ and selected on LB plates containing 50 μg/mL apramycin and 50 μg/mL kanamycin.

    Spectrophotometry:

    Article Title: Chromosome-Specific DNA Repeats: Rapid Identification in Silico and Validation Using Fluorescence in Situ Hybridization
    Article Snippet: Fluorescence in situ Hybridization Individual BAC clones were grown overnight in up to 10 mL of Luria broth (LB) medium containing 12.5 μg/mL chloramphenicol (Sigma, St. Louis, MO, USA), and the DNA was isolated using a ZR BAC DNA Miniprep Kit (Zymo Research, Irvine, CA, USA). .. The isolation of high molecular weight BAC DNAs was confirmed on 1% agarose gels and quantitated by spectrophotometry (Nanodrop 2000, Thermo Scientific, Wilmington, DE, USA).

    FLAG-tag:

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: A sequence encoding the FLAG tag was added to the N-terminus of MCM3. .. The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research).

    Alkaline Lysis:

    Article Title: Bioinformatics Tools Allow Targeted Selection of Chromosome Enumeration Probes and Aneuploidy Detection
    Article Snippet: .. DNA Probe Preparation The BAC DNAs were extracted from overnight cultures following an alkaline lysis protocol ( ) or using a ZR BAC DNA Miniprep Kit (Zymo Research; Irvine, CA). .. The isolation of high molecular weight BAC DNAs was confirmed on 1% agarose gels and quantitated by Hoechst fluorometry using a Hoefer TK 100 instrument (Hoefer; South San Francisco, CA).

    BAC Assay:

    Article Title: Multiple organ gigantism caused by mutation in VmPPD gene in blackgram (Vigna mungo)
    Article Snippet: .. After screening, the BAC DNA of the clone VmR48p23 was extracted using ZR BAC DNA Miniprep Kit (Zymo Research). .. The extracted DNA was sheared into ~3 kb fragments, cloned into pUC118 vector, sequenced with BigDye Terminator (Thermo Fisher Scientific) and ABI3730 (Thermo Fisher Scientific), assembled by CLC Genomic Workbench (QIAGEN) and annotated manually.

    Article Title: Chromosome-Specific DNA Repeats: Rapid Identification in Silico and Validation Using Fluorescence in Situ Hybridization
    Article Snippet: .. Fluorescence in situ Hybridization Individual BAC clones were grown overnight in up to 10 mL of Luria broth (LB) medium containing 12.5 μg/mL chloramphenicol (Sigma, St. Louis, MO, USA), and the DNA was isolated using a ZR BAC DNA Miniprep Kit (Zymo Research, Irvine, CA, USA). .. For the preparation of DNA pools, clones were grown individually and pooled prior to DNA isolation.

    Article Title: Establishment of an Efficient and Flexible Genetic Manipulation Platform Based on a Fosmid Library for Rapid Generation of Recombinant Pseudorabies Virus
    Article Snippet: .. The fosmids were extracted using ZR BAC DNA Miniprep Kit (Zymo Research). .. Fosmid end-sequencing was performed using pCC1FOS sequencing primers.

    Article Title: Colibactin assembly line enzymes use S-adenosylmethionine to build a cyclopropane ring
    Article Snippet: .. The mutant BACs were isolated from these colonies using ZR BAC DNA Miniprep Kit (Zymo research), and gene disruption was verified by PCR amplification of knockout regions. .. To generate an E. coli mutant strain expressing ClbA, ClbB, ClbC, and ClbN, E. coli DH10B were electroporated with mutant BACpks ΔclbDtoM ΔclbOtoQ and selected on LB plates containing 50 μg/mL apramycin and 50 μg/mL kanamycin.

    Article Title: A human monoclonal antibody that specifically binds and inhibits the staphylococcal complement inhibitor protein SCIN
    Article Snippet: .. Bacterial chromosomal DNA was isolated using the ZR BAC DNA Miniprep Kit (Zymo Reasearch Corporation, USA) following the manufacturer's protocol. ..

    Article Title: A PECTIN METHYLESTERASE gene at the maize Ga1 locus confers male function in unilateral cross-incompatibility
    Article Snippet: .. A 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA). ..

    Article Title: LANA oligomeric architecture is essential for KSHV nuclear body formation and viral genome maintenance during latency
    Article Snippet: .. Briefly, BAC16 DNA was isolated from 2ml bacterial culture using a ZR BAC DNA Miniprep Kit (Zymo Research) and resuspended in 30 μl of RNase-free water. iSLK cells were seeded at 2x105 cells/well of a 6-well plate. .. On the next day 15 μl of BAC DNA was mixed with transfection solution and added to the cells following by transfection kit’s according to manufacturer’s protocol.

    Article Title: Escape of X-linked miRNA genes from meiotic sex chromosome inactivation
    Article Snippet: .. BAC/fosmid DNA was isolated using the ZR BAC DNA miniprep Kit (ZymoResearch) according to the manufacturer's protocol. .. DNA concentration and quality were characterized by agarose gel electrophoresis and analysis on a BioRad Nanodrop.

    Article Title: Wheat Ms2 encodes for an orphan protein that confers male sterility in grass species
    Article Snippet: .. In brief, a 384-well stock was duplicated and pooled to extract a primary DNA pool using the ZR BAC DNA Miniprep Kit (Zymo Research Corporation, Irvine, CA, USA). ..

    Article Title: TRAIP is a master regulator of DNA interstrand cross-link repair
    Article Snippet: .. The bacmid encoding the CMG complex was obtained by electroporating pGC187 into DH10EMBacY (Geneva Biotech) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research). .. CMG baculovirus was generated by transfecting pGC187 bacmid into Sf9 cells (Expression Systems) for 96 hr, and amplifying the virus in 2 rounds of 96 hr each.

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: .. The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048). .. Baculovirus encoding GINS was amplified in three stages (P1, P2, and P3) in Sf9 cells (Expression Systems Cat# 94–001S) and GINS expression levels were monitored via western blots.

    Article Title: Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA
    Article Snippet: .. BACs to be used as templates for long PCR or for screening by restriction enzyme digestion were purified from 4 ml overnight cultures of E. coli DH10B using the ZR BAC DNA Miniprep Kit (Zymo Research). .. BACs required for direct genome sequencing were purified from 500 ml cultures using the Large-construct kit (Qiagen).

    Article Title: Bioinformatics Tools Allow Targeted Selection of Chromosome Enumeration Probes and Aneuploidy Detection
    Article Snippet: .. DNA Probe Preparation The BAC DNAs were extracted from overnight cultures following an alkaline lysis protocol ( ) or using a ZR BAC DNA Miniprep Kit (Zymo Research; Irvine, CA). .. The isolation of high molecular weight BAC DNAs was confirmed on 1% agarose gels and quantitated by Hoechst fluorometry using a Hoefer TK 100 instrument (Hoefer; South San Francisco, CA).

    Lysis:

    Article Title: The CMG helicase bypasses DNA-protein cross-links to facilitate their repair
    Article Snippet: The bacmid encoding the GINS complex was obtained by electroporating pGC128 into DH10EMBacY (or DH10Bac) electro-competent cells and purified using ZR BAC DNA miniprep kit (Zymo Research Cat# D4048). .. Cells harvested 48 hrs post-infection were pelleted at 500 × g for 15min and re-suspended in a final volume of 50mL in GINS Lysis Buffer (GLB) containing 20mM Tris-HCl pH8.0, 5% glycerol, 500mM NaCl, 20mM Imidazole, 1mM DTT, 1mM PMSF, EDTA-free cOmplete protease inhibitor cocktail (Roche Cat # 11873580001).

    Chick Chorioallantoic Membrane Assay:

    Article Title: Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA
    Article Snippet: All BACs were modified and maintained in E. coli DH10B cells (Invitrogen) grown at 37°C in LB medium containing chloramphenicol (Cam, 15 μg/ml). .. BACs to be used as templates for long PCR or for screening by restriction enzyme digestion were purified from 4 ml overnight cultures of E. coli DH10B using the ZR BAC DNA Miniprep Kit (Zymo Research).

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    Zymo Research bac dna miniprep kit
    Screen dump of the UCSC Genome Browser (GoldenPath) display showing the <t>BAC</t> ends mapped to the targeted region of chromosome 2: 92,269,115 bp to 92,335,426 bp. The alignment BAC clone end sequences with the draft sequence of the human genome places BAC RP11-100H17 (arrow) in a region comprised entirely of satellite <t>DNA,</t> while BAC RP11-469P16 (arrowhead) is predicted to contain a cluster of long interspersed repeated DNA sequences (LINEs)(circled).
    Bac Dna Miniprep Kit, supplied by Zymo Research, used in various techniques. Bioz Stars score: 95/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Screen dump of the UCSC Genome Browser (GoldenPath) display showing the BAC ends mapped to the targeted region of chromosome 2: 92,269,115 bp to 92,335,426 bp. The alignment BAC clone end sequences with the draft sequence of the human genome places BAC RP11-100H17 (arrow) in a region comprised entirely of satellite DNA, while BAC RP11-469P16 (arrowhead) is predicted to contain a cluster of long interspersed repeated DNA sequences (LINEs)(circled).

    Journal: Current Genomics

    Article Title: Bioinformatic Tools Identify Chromosome-Specific DNA Probes and Facilitate Risk Assessment by Detecting Aneusomies in Extra-embryonic Tissues

    doi: 10.2174/138920212802510510

    Figure Lengend Snippet: Screen dump of the UCSC Genome Browser (GoldenPath) display showing the BAC ends mapped to the targeted region of chromosome 2: 92,269,115 bp to 92,335,426 bp. The alignment BAC clone end sequences with the draft sequence of the human genome places BAC RP11-100H17 (arrow) in a region comprised entirely of satellite DNA, while BAC RP11-469P16 (arrowhead) is predicted to contain a cluster of long interspersed repeated DNA sequences (LINEs)(circled).

    Article Snippet: In typical experiments, the BAC DNAs are extracted from overnight cultures following an alkaline lysis protocol [ ] or using a BAC DNA miniprep kit (Zymo Research; Irvine, CA).

    Techniques: BAC Assay, Sequencing

    Screen dump of the UCSC Genome Browser display showing the BAC ends mapped to the region chromosome 4: 49,093,534 bp to 49,183,369 bp. The alignment BAC clone end sequences with the draft sequence of the human genome places BAC RP11-360M1 (high-lighted) in a region comprised almost entirely of satellite DNA and completely free of interspersed repeated DNA sequences such as short interspersed repeats (SINEs) or LINEs.

    Journal: Current Genomics

    Article Title: Bioinformatic Tools Identify Chromosome-Specific DNA Probes and Facilitate Risk Assessment by Detecting Aneusomies in Extra-embryonic Tissues

    doi: 10.2174/138920212802510510

    Figure Lengend Snippet: Screen dump of the UCSC Genome Browser display showing the BAC ends mapped to the region chromosome 4: 49,093,534 bp to 49,183,369 bp. The alignment BAC clone end sequences with the draft sequence of the human genome places BAC RP11-360M1 (high-lighted) in a region comprised almost entirely of satellite DNA and completely free of interspersed repeated DNA sequences such as short interspersed repeats (SINEs) or LINEs.

    Article Snippet: In typical experiments, the BAC DNAs are extracted from overnight cultures following an alkaline lysis protocol [ ] or using a BAC DNA miniprep kit (Zymo Research; Irvine, CA).

    Techniques: BAC Assay, Sequencing

    In situ hybridization analysis of DNA probes prepared from BAC clones. The BAC clones RP11-294C12 and RP11-242E13 hybridized to metaphase spreads prepared from short term cultures of human lymphocytes showed specific hybridization to the target regions on the X (arrowhead) and Y (arrow) chromosome, respectively. A) Schematic representation of the FISH target regions on the X and Y chromosome. B) Hybridization of both probes to metaphase chromosomes. C) Hybridization signals in diploid interphase cell nuclei. (Bars = 10 µm).

    Journal: Current Genomics

    Article Title: Bioinformatic Tools Identify Chromosome-Specific DNA Probes and Facilitate Risk Assessment by Detecting Aneusomies in Extra-embryonic Tissues

    doi: 10.2174/138920212802510510

    Figure Lengend Snippet: In situ hybridization analysis of DNA probes prepared from BAC clones. The BAC clones RP11-294C12 and RP11-242E13 hybridized to metaphase spreads prepared from short term cultures of human lymphocytes showed specific hybridization to the target regions on the X (arrowhead) and Y (arrow) chromosome, respectively. A) Schematic representation of the FISH target regions on the X and Y chromosome. B) Hybridization of both probes to metaphase chromosomes. C) Hybridization signals in diploid interphase cell nuclei. (Bars = 10 µm).

    Article Snippet: In typical experiments, the BAC DNAs are extracted from overnight cultures following an alkaline lysis protocol [ ] or using a BAC DNA miniprep kit (Zymo Research; Irvine, CA).

    Techniques: In Situ Hybridization, BAC Assay, Clone Assay, Hybridization, Fluorescence In Situ Hybridization

    FISH performance of BAC-derived DNA probes targeting the centromeric heterochromatin of chromosome 2. A-B) A DNA probe prepared from BAC clone RP11-469P16 shows multiple signals in normal interphase cell nuclei ( A ) or on metaphase spreads ( B ). Arrows in B ) point at the target region on chromosome 2. C ) A DNA probe prepared from BAC clone RP11-100H17 binds exclusively to the chromosome 2-specific target region (arrows). (Bars = 10 µm).

    Journal: Current Genomics

    Article Title: Bioinformatic Tools Identify Chromosome-Specific DNA Probes and Facilitate Risk Assessment by Detecting Aneusomies in Extra-embryonic Tissues

    doi: 10.2174/138920212802510510

    Figure Lengend Snippet: FISH performance of BAC-derived DNA probes targeting the centromeric heterochromatin of chromosome 2. A-B) A DNA probe prepared from BAC clone RP11-469P16 shows multiple signals in normal interphase cell nuclei ( A ) or on metaphase spreads ( B ). Arrows in B ) point at the target region on chromosome 2. C ) A DNA probe prepared from BAC clone RP11-100H17 binds exclusively to the chromosome 2-specific target region (arrows). (Bars = 10 µm).

    Article Snippet: In typical experiments, the BAC DNAs are extracted from overnight cultures following an alkaline lysis protocol [ ] or using a BAC DNA miniprep kit (Zymo Research; Irvine, CA).

    Techniques: Fluorescence In Situ Hybridization, BAC Assay, Derivative Assay

    Culture genotype of chromosome 10 in a PTC cell line. Chromosome numbers and signal counts of the chromosome 10-specific DNA probe prepared from BAC RP11-168p20 hybridized to cell line S48TK18A6 (scored by three independent scorers).

    Journal: Journal of Histochemistry and Cytochemistry

    Article Title: Bioinformatics Tools Allow Targeted Selection of Chromosome Enumeration Probes and Aneuploidy Detection

    doi: 10.1369/0022155412470955

    Figure Lengend Snippet: Culture genotype of chromosome 10 in a PTC cell line. Chromosome numbers and signal counts of the chromosome 10-specific DNA probe prepared from BAC RP11-168p20 hybridized to cell line S48TK18A6 (scored by three independent scorers).

    Article Snippet: DNA Probe Preparation The BAC DNAs were extracted from overnight cultures following an alkaline lysis protocol ( ) or using a ZR BAC DNA Miniprep Kit (Zymo Research; Irvine, CA).

    Techniques: BAC Assay

    In situ hybridization results for selected DNA probes. Red shaded areas on the idiograms in (A–B) and (F–G) indicate the hybridization target sites. (A) In situ hybridization using fluorescent probes for the highly repeated satellite III DNA segment (flanked by the PCR primers WYR2 and WYR4) shows bright signals on the long arm of the Y chromosome. (B) Combining biotinylated in situ hybridization probes for the Y chromosome-specific DNA segment (large signal) with probes for X chromosome-specific DNA flanked by the PCR primers WXR1 and WXR2 (small signal) allows chromosome identification based on the size of the signal. (C) A digoxigenin-labeled DNA probe prepared from BAC RP11-348g24 resulted in specific signals on the X chromosome in the presence of a significant amount of cross-hybridization to other chromosomes in metaphase spreads. High level of cross-hybridization was also evident in interphase nuclei. (D) In contrast to the result shown in (C), a probe prepared from BAC RP11-294c12 bound almost exclusively to the centromeric heterochromatin of the human X chromosome. (E) Combining differentially labeled DNA probes in a single hybridization experiment, the Y chromosomal target appears in green, whereas the X chromosomal centromeric repeat DNA is shown in red. (F) In situ hybridization of an autosome-targeting DNA probe prepared from BAC clone RP11-96f8 showed multiple signals and a high level of cross-hybridization in the interphase cell nuclei. (G) The biotinylated probe prepared from BAC RP11-168p20 exhibited strong signals on both homologues of chromosome 10. (H) A combination of three differently labeled probes was hybridized simultaneously. Chromosome 10 signals are shown in red; X chromosomal and Y chromosomal signals are shown in green and blue, respectively. (I–K) The same probe as in (G) (BAC RP11-168p20) was hybridized to the cell line S48TK18A6. Multiple signals indicate chromosomal abnormalities. (I) shows DAPI picture, (J) shows green avidin-FITC signal, and (K) shows the superposition of both. (L–N) A chromosome X probe (BAC clone RP11-294c12) labeled with biotin/avidin-FITC, a Cy-5 labeled chromosome Y probe (RP11-242e13), and a chromosome 10-specific probe prepared from BAC RP11-168p20 (labeled with Spectral Orange-dUTP) were hybridized onto a deparaffinized 12-week placenta tissue section. (L) The chromosome 10 probe (red signals) and bound chromosome X probe (green signals) in this female placental tissue section. (M–N) In our triple probe FISH experiment, the chromosome 10 probe is represented by orange signals, the chromosome X probe by green signals, and the chromosome Y probe by red signals in the male cell nuclei. (Size marker bars indicate 10 µm.)

    Journal: Journal of Histochemistry and Cytochemistry

    Article Title: Bioinformatics Tools Allow Targeted Selection of Chromosome Enumeration Probes and Aneuploidy Detection

    doi: 10.1369/0022155412470955

    Figure Lengend Snippet: In situ hybridization results for selected DNA probes. Red shaded areas on the idiograms in (A–B) and (F–G) indicate the hybridization target sites. (A) In situ hybridization using fluorescent probes for the highly repeated satellite III DNA segment (flanked by the PCR primers WYR2 and WYR4) shows bright signals on the long arm of the Y chromosome. (B) Combining biotinylated in situ hybridization probes for the Y chromosome-specific DNA segment (large signal) with probes for X chromosome-specific DNA flanked by the PCR primers WXR1 and WXR2 (small signal) allows chromosome identification based on the size of the signal. (C) A digoxigenin-labeled DNA probe prepared from BAC RP11-348g24 resulted in specific signals on the X chromosome in the presence of a significant amount of cross-hybridization to other chromosomes in metaphase spreads. High level of cross-hybridization was also evident in interphase nuclei. (D) In contrast to the result shown in (C), a probe prepared from BAC RP11-294c12 bound almost exclusively to the centromeric heterochromatin of the human X chromosome. (E) Combining differentially labeled DNA probes in a single hybridization experiment, the Y chromosomal target appears in green, whereas the X chromosomal centromeric repeat DNA is shown in red. (F) In situ hybridization of an autosome-targeting DNA probe prepared from BAC clone RP11-96f8 showed multiple signals and a high level of cross-hybridization in the interphase cell nuclei. (G) The biotinylated probe prepared from BAC RP11-168p20 exhibited strong signals on both homologues of chromosome 10. (H) A combination of three differently labeled probes was hybridized simultaneously. Chromosome 10 signals are shown in red; X chromosomal and Y chromosomal signals are shown in green and blue, respectively. (I–K) The same probe as in (G) (BAC RP11-168p20) was hybridized to the cell line S48TK18A6. Multiple signals indicate chromosomal abnormalities. (I) shows DAPI picture, (J) shows green avidin-FITC signal, and (K) shows the superposition of both. (L–N) A chromosome X probe (BAC clone RP11-294c12) labeled with biotin/avidin-FITC, a Cy-5 labeled chromosome Y probe (RP11-242e13), and a chromosome 10-specific probe prepared from BAC RP11-168p20 (labeled with Spectral Orange-dUTP) were hybridized onto a deparaffinized 12-week placenta tissue section. (L) The chromosome 10 probe (red signals) and bound chromosome X probe (green signals) in this female placental tissue section. (M–N) In our triple probe FISH experiment, the chromosome 10 probe is represented by orange signals, the chromosome X probe by green signals, and the chromosome Y probe by red signals in the male cell nuclei. (Size marker bars indicate 10 µm.)

    Article Snippet: DNA Probe Preparation The BAC DNAs were extracted from overnight cultures following an alkaline lysis protocol ( ) or using a ZR BAC DNA Miniprep Kit (Zymo Research; Irvine, CA).

    Techniques: In Situ Hybridization, Hybridization, Polymerase Chain Reaction, Labeling, BAC Assay, Avidin-Biotin Assay, Fluorescence In Situ Hybridization, Marker