molecular genetics recombinant dna amp genomic technology

... using recombinant DNA technology was human insulin, developed by Genentech and Licensed by Eli Lilly and Company  Applications of recombinant biotechnology  Recombinant human insulin  Recombinant ... successful production and intracellular replication of recombinant DNA appeared in 1972 and 1973 Stanford University applied for a US patent on recombinant DNA in 1974, listing the inventors as Stanley ... Self-replicating  Double stranded  Circular DNA  Independent extrachromosomal DNA in  Ampicillin bacteria 1kb-500kb  Kanamycin  Origin of DNA replication, ori  High (10-100) & low copy...

Ngày tải lên: 23/03/2014, 22:20

... DNA pieces are joined in vitro to form recombinant molecules • Generate sticky ends on the DNA, e.g with restriction endonucleases • Tie DNA molecules from different sources together with DNA ... CTTAA G Recombinant molecules G AATTC CTTAA G GAATTC CTTAAG GAATTC CTTAAG Parental molecules DNA ligase covalently joins two DNA molecules • Uses ATP or NADH to provide energy to seal nicks DNA ligase ... cycles, the amplification is approximately [2exp(n-1)]-2 • After 21 cycles, the fragment has been amplified about a million-fold • E.g a sample with 0.1 pg of the target fragment can be amplified...

Ngày tải lên: 13/03/2014, 18:39

... steps used to clone DNA?     Isolate DNA from an organism Cut the organismal DNA and the vector with restriction enzymes making recombinant DNA Introduce the recombinant DNA into a host Restriction ... of DNA Using DNA probes to screen plasmid genomic libraries for specific DNA sequences PowerPoint® Layered Art Figure 8.11 Using DNA probes to screen plasmid genomic libraries for specific DNA ... to produce DNA fragments of appropriate size for constructing a genomic library cDNA Libraries are best for eukaryotes Figure 8.8 The synthesis of double-stranded complementary DNA (cDNA) from...

Ngày tải lên: 13/03/2014, 19:33

... Isoplant DNA extraction Kit (Takara Bio Inc., Shiga, Japan) PCR was performed using KOD DNA polymerase (Toyobo, Osaka, Japan) and DNA AMPLIFIER MIR-D40 (Sanyo, Osaka, Japan) To amplify the DNA fragments ... Matsuo et al DNA isolation and PCR amplification The x-5 gliadin gene was cloned from a Japanese soft wheat cultivar, Norin 61 (Shimane Agricultural Experiment Station) Total genomic DNA was isolated ... sequence tags (ESTs) A high-fidelity DNA polymerase was used to reduce the risk of introducing errors into the sequence Amplification of genes from wheat genomic DNA produced two products designated...

Ngày tải lên: 20/02/2014, 01:20

... screening with molecular testing has helped to clarify what might have first seemed confusing [1] Three CCM genes have been mapped and identified in the recent years These molecular genetics data ... exons of the three CCM genes and search for genomic rearrangements using cDNA and ⁄ or quantitative multiplex PCR such as multiplex ligation-dependent probe amplification in Caucasian non-Hispano-American ... Authors Journal compilation ª 2010 FEBS 1073 CCM molecular genetics F Riant et al References Labauge P, Denier C, Bergametti F & Tournier-Lasserve E (2007) Genetics of cavernous angiomas Lancet Neurol...

Ngày tải lên: 15/03/2014, 10:20

... Craniosynostoses Molecular Genetics, Principles of Diagnosis, and Treatment Monographs in Human Genetics Vol 19 Series Editor Michael Schmid Würzburg Craniosynostoses Molecular Genetics, Principles ... Chapter Saethre-Chotzen Syndrome: Clinical and Molecular Genetic Aspects Kress, W.; Collmann, H (Würzburg) Chapter 10 Craniofrontonasal Syndrome: Molecular Genetics, EFNB1 Mutations and the Concept ... great pleasure to introduce volume 19 of the book series Monographs in Human Genetics entitled ‘Craniosynostoses: Molecular Genetics, Principles of Diagnosis and Treatment’ The initial idea for this...

Ngày tải lên: 28/03/2014, 22:20

... in the sample of interest are identified based on the signal emitted from the array Nylon filter-based arrays are gridded with DNA fragments, usually derived from cDNAs PCR-amplified cDNAs are ... aspects in molecular genetics such as functional genomics, ESTs, and SSRs, and the profound case studies addressing temperate and tropical trees.” Prof Dr Reiner Finkeldey Chair of Forest Genetics ... Vidhyaskdaran Molecular Genetics and Breeding of Forest Trees edited by Sandeep Kumar and Matthias Fladung Testing of Genetically Modified Organisms in Foods edited by Farid E Ahmed Molecular Genetics...

Ngày tải lên: 03/04/2014, 12:10

... obligatory elongation of a DNA strand only at the 3' end is an essential feature of DNA replication Chapter Introduction to Molecular Genetics and Genomics (A) (B) Parent molecule of DNA 5’ T A 3’ ACGCT ... This approach to the study of genetics is often referred to as classical genetics, or organismic or morphological genetics Given the advances of molecular, or modern, genetics, it is possible to ... book we include many examples showing how molecular and classical genetics can be used in combination to enhance the power of genetic analysis The foundation of genetics as a molecular science dates...

Ngày tải lên: 08/04/2014, 13:07

... See recombinant DNA technologies Recombinant DNA molecules: A combination of DNA molecules of different origin that are joined using recombinant DNA technologies 40 Recombinant DNA technologies: ... ancestor In recombinant DNA technology, the use of DNA manipulation procedures to produce multiple copies of a single gene or segment of DNA is referred to as cloning DNA Cloning vector: DNA molecule ... Clones Chromosomal DNA Vector DNA 21 PCR (in vitro DNA amplification) Described as being to genes what Gutenberg’s printing press was to the written word, PCR can amplify a desired DNA sequence of...

Ngày tải lên: 11/04/2014, 07:15

... cell genetics with recombinant DNA and gene transfer technology has resulted in new approaches especially useful for the study of mutant cells This volume illustrates how special techniques in molecular ... cells in culture Basic protocols for the manipulation of recombinant DNA can be found in other Methods in Enzymology volumes: Recombinant DNA, Parts A - F , Volumes 68, 100, 101,153, 154, and 155 ... York, 1985 M M Gottesman, in "Molecular Cell Genetics" (M M Gottesrnan, ed.), p 139 Wiley, New York, 1985 M J Siciliano, R L Stallings, and G M Adair, in "Molecular Cell Genetics" (M M Gottesman,...

Ngày tải lên: 11/04/2014, 09:52

... site, to help verify amplification of the correct product An intron sequence, to distinguish genomic amplification product from those amplified from cDNA or contaminating DNA (see Section 6) A ... of too many cycles of PCR, which favors the amphficauon of mismatched or nonspectfic DNA products For amplification of a smglecopy gene from genomic DNA, 35 cycles should be enough, but more cycles ... Strand cDNA s PCR Ampl If led cDNA CDNA w x l Dldeoxynucleotlde Seauenclng Seauenclng Primer Template Excess of Single Strands Seauence Pig Schematic dmgram of the strategy for direct DNA sequencing...

Ngày tải lên: 11/04/2014, 10:14

... host cell population have no recombinant DNA replicating in them This selection or screening for desired recombinants provides a route to recovery of the recombinant DNA of interest in pure form ... cloning of yeast DNA, or certain prokaryotic DNAs, in yeast A vehicle for recombinant DNA and a transformation procedure are also available for Staphylococcus aureus.S2 Many recombinants containing ... and can discover what portions of the DNA are most closely related to that phenotype Conclusion Recombinant DNA methods have become a mainstay of molecular genetics They are also contributing to...

Ngày tải lên: 11/04/2014, 10:25

... degree of purity of the DNA substrate Improperly prepared DNA samples will be cleaved poorly, if at all, producing partially digested DNA In addition to DNA purity, other DNA- associated parameters ... (80/zg/ml) of lambda DNA in a 15-hr reaction 16 This phenomenon may be attributed to the viscosity produced by high concentrations of high molecular weight DNA (e.g., lambda DNA) , which can inhibit ... plasmid or viral DNAs completely than for linear DNA A comparison of the relative cleavage efficiencies for several supercoiled and linear DNAs are presented in Table V If a supercoiled DNA (e.g.,...

Ngày tải lên: 11/04/2014, 10:25

... CLONING GENES Vector DNA Bamred + ),,+ Vector contains lambda red and gamma genes on 17 Kb Bam H1 fragment Barn Digest phage DNA with BamH1 High Molecular Weight Eukaryotic DNA Digest DNA with Bam H1, ... of nematode DNA fragments produce recombinant phages that are detected by plating on Q359 The yield of recombinants is linear with the amount of nematode DNA added as long as the DNA concentration ... of insert DNA to vector DNA (0.5/~g insert DNA per 1.0/~g of vector) The yield of recombinants in this experiment ranged from 2.4 x 10~ to 5.4 x 105 per microgram of 15-20 kb nematode DNA This...

Ngày tải lên: 11/04/2014, 10:26

... of ssDNA Selective Packaging of the (Recombinant) DNA Strands of pKUN The production of ss recombinant DNA with this system in many respects resembles that for obtaining recombinant M13 DNA 1°,12Therefore ... ss phage DNA and the other ss plasmid DNA (see Fig 3A) Sometimes you might also see traces of high -molecular- weight DNA In case of packaging by IKe-9 another, fast migrating, (miniphage DNA) band ... prokaryotic origin Some examples of packaging of ss recombinant plasmid DNA are shown in Fig 3A The level and ratio of ss phage DNA (upper band in Fig 3A) to ss plasmid DNA (lower band) vary for...

Ngày tải lên: 11/04/2014, 10:26

... formation of high molecular weight DNA by base-pairing between dG-tails, the yield of linker DNA from agarose gels is generally poor cDNA Cloning Steps in the construction of cDNA-plasmid recombinants ... OF ~ DNA POLYMERASE1 AND DNA UGASE J /H~m DIGESTION Xho[8BamHl GGc~'~-~ "~" Hk'~dH] 'P~ tl~ A TAA FIG Enzymatic steps in the construction of pcD-cDNA recombinants The designations of the DNA segments ... RNA strand is converted to DNA by nicktranslation repair catalyzed by Escherichia coli DNA polymerase I, RNase H, and DNA ligase The end product, a closed circular cDNA recombinant, is transfected...

Ngày tải lên: 11/04/2014, 10:26

... Site-specific DNA methylases (S-adenosylmethionine : DNA methyltransferases) have a variety of uses in DNA analytical procedures Methylases may be used as probes for DNA conformation,L2 in recombinant DNA ... sequence specificities Materials Restriction endonucleases, purified DNA methylases, T4 DNA ligase, unmethylated phage h DNA, ~bX174 RF I DNA, and 5'-phosphorylated oligonucleotides were obtained from ... and selective DNA methylase assay early in many DNA methylase purifications For example, the M.MbolI methylase must be assayed in the presence of M.MboI and at least three other DNA methylases...

Ngày tải lên: 11/04/2014, 10:26

... of DNA are captured from large sample volumes, e.g., 10 ng of DNA from 0.5 ml of sample; (5) relatively large amounts of DNA are captured from small volumes, e.g., /~g of DNA from 30 /zl of sample; ... high molecular weight DNA molecules, socalled megabase DNA (Mbp DNA) For various reasons, plant molecular biologists have been slow to develop specific protocols suitable for preparing Mbp DNA ... (50), Department of Human Genetics and Molecular Biology, Collaborative Research, Inc., Waltham, Massachusetts 02154 ADRIENNE P SMYTH (50), Department of Human Genetics and Molecular Biology, Collaborative...

Ngày tải lên: 11/04/2014, 10:26

... Cloning of DNA Fragments Any of several reliable methods can be used to prepare DNA/ cDNA for cloning in pIFFS For direct in-frame cloning in pIFF8 (prepared as above) the foreign DNA fragment ... not will remain Lac + Examples Selection of Open Reading Frames in DNA/ cDNA The pIFF series of vectors, and in particular pIFF8, are recent vector constructions and examples on experimental applications ... expression [4] M D R I GENE IN MAMMALIAN EXPRESSION VECTORS 45 DNA Detection Transfected MDR1 DNA can be detected by isolating genomic DNA from multidrug-resistant cells and performing Southern analysis...

Ngày tải lên: 11/04/2014, 10:26

... (1991) FIG Sample sequencing ladders obtained from PCR-amplified ssDNA (A-C) Amplifications from Mus spretus genomic DNA; (D-F) amplifications from Drosophila melanogaster genomic DNA (A and B) ... product directly from genomic DNA relies on having an efficient method for sequencing the amplified DNA Several approaches have been taken for sequencing PCR-amplified DNA The ease and reproducibility ... Acad Sci U.S.A 86, 6196 (1989) a 91011 121314 dsDNA b 91011121314 dsDNA ssDNA =~ C dsDNA =~ 91011121314 [1] SEQUENCING OF in Vitro AMPLIFIED DNA 50/1 50/2 50/3 50/4 5o/5 1150 2/50 3/50 4•50...

Ngày tải lên: 11/04/2014, 10:27