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cleavage site e.g. BamHI. Both the cDNA and the linker have blunt ends which can be ligated together using a high concentration of T4 DNA ligase. Then sticky ends are produced in the cDNA molecule by cleaving the cDNA ends (which now have linkers with an incorporated site) with the appropriate
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cDNA libraries are commonly used when reproducing eukaryotic genomes, as the amount of information is reduced to remove the large numbers of non-coding regions from the library. cDNA libraries are used to express eukaryotic genes in prokaryotes. Prokaryotes do not have introns in their DNA and
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Firstly, mRNA template needs to be isolated for the creation of cDNA libraries. Since mRNA only contains exons, the integrity of the isolated mRNA should be considered so that the protein encoded can still be produced. Isolated mRNA should range from 500 bp to 8 kb. Several methods exist for
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I is then added, the cleaved RNA acts as a primer the DNA polymerase I can identify and initiate replacement of RNA nucleotides with those of DNA. This is provided by the sscDNA itself by coiling on itself at the 3' end, generating a
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The cloned bacteria are then selected, commonly through the use of antibiotic selection. Once selected, stocks of the bacteria are created which can later be grown and sequenced to compile the cDNA library.
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to identify genes based on the encoded protein's function. When studying eukaryotic DNA, expression libraries are constructed using complementary DNA (cDNA) to help ensure the insert is truly a gene.
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therefore do not possess any enzymes that can cut it out during transcription process. cDNA does not have introns and therefore can be expressed in prokaryotic cells. cDNA libraries are most useful in
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and can be readily expressed in a bacterial cell. While information in cDNA libraries is a powerful and useful tool since gene products are easily identified, the libraries lack information about
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primer (a short sequence of deoxy-thymidine nucleotides) is bound to the poly-A tail of the RNA. The primer is required to initiate DNA synthesis by the enzyme
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enzyme is used to cleave the backbone of the mRNA and generate free 3'-OH groups, which is important for the replacement of mRNA with DNA.
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and therefore contains only the expressed genes of an organism. Similarly, tissue-specific cDNA libraries can be produced. In
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cDNA molecules can be cloned by using restriction site linkers. Linkers are short, double stranded pieces of DNA (
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provide more detailed information about the organism, but are more resource-intensive to generate and keep.
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where the additional genomic information is of less use. Additionally, cDNA libraries are frequently used in
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site for reverse transcription. This has the problem that not all transcripts, such as those for the
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cDNA library || How cDNA library is constructed? || What are DNA libraries used for?
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cDNA library lacks the non-coding and regulatory elements found in genomic DNA.
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507:. Pazdernik, Nanette Jean. Amsterdam: Academic Press/Elsevier.
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396:"cDNA Libraries - an overview | ScienceDirect Topics"
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Biotechnology : applying the genetic revolution
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553:Functional Annotation of the Mouse database
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447:"Complementary DNA Libraries: An Overview"
109:Learn how and when to remove this message
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189:from a eukaryotic cell with the use of
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342:oligodeoxyribonucleotide
451:Molecular Biotechnology
445:Ying, Shao-Yao (2004).
367:host cell for cloning.
230:purifying RNA such as
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330:Genomic DNA libraries
256:reverse transcriptase
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43:improve this article
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236:column purification
193:. In eukaryotes, a
167:genomic DNA library
16:Type of DNA library
377:Functional cloning
318:functional cloning
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578:Molecular biology
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41:Please help
36:verification
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219:poly-A tail
217:, encode a
572:Categories
427:2024-02-19
405:2024-02-19
383:References
359:sticky end
289:DNA ligase
147:eukaryotic
69:newspapers
531:cite book
523:226038060
471:1073-6085
159:enhancers
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371:See also
297:plasmids
280:nuclease
252:oligo-dT
99:May 2008
364:E. coli
355:plasmid
260:RNAse H
215:histone
163:introns
155:introns
151:spliced
143:nucleus
135:library
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211:primer
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201:from
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76:books
537:link
519:OCLC
509:ISBN
475:PMID
467:ISSN
287:and
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205:and
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199:mRNA
187:mRNA
139:mRNA
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