GENE SYNTHESIS & SEQUENCING

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METHOD TO INCREASE THE CONCENTRATION OF OLIGONUCLEOTIDES TO LEVELS SUITABLE FOR GENE SYNTHESIS:

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On the 30 -stop of each oligonucleotide produced at the array, there's a not unusual place priming series accompanied with the aid of using a nicking endonuclease popularity site. The array-synthesized oligonucleotides function the template strand for a primer extension manner that employs the not unusual place priming series that became covered into the array-connected oligonucleotide pool.

 

Following primer extension, a nicking endonuclease withinside the response combination removes the not unusual place primer sequences, ensuing in a single-stranded DNA oligonucleotide similar to every oligonucleotide produced at the array. 

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This manner amplifies the array-synthesized oligonucleotide pool to growth the awareness of the oligonucleotides to degrees appropriate for gene synthesis.

 

ANOTHER METHOD:

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Another technique advanced through Kosuri et al. addresses the oligonucleotide awareness and pool complexity problems through including “bar-coded” priming sequences into every of the oligonucleotides produced at the array. The oligonucleotides important to fabricate a given synthon are bordered on each ends through units of preset primer sequences on this technique.

 

The amplification of the array-produced oligonucleotide pool, which has been cleaved from the chip floor in the course of synthesis, is enabled through the outermost priming sequences. 

This technique allows a unmarried set of particular primers to expand the entire oligonucleotide pool, permitting part of the pool to be utilized for destiny meeting or archiving.

 

A series of flanking primer-binding sequences inboard of the pool amplification primers lets in for selective amplification of meeting sub swimming pools from the synthesis pool. Each wonderful synthon could be assigned a wonderful set of flanking subpool primer sequences, permitting the oligo nucleotides required to assemble any precise synthon to be preferentially amplified from the synthesis pool for destiny meet. 

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Following sub pool oligonucleotide amplification, the priming sequences are removed through restrict digestion of the amplified dsDNA oligonucleotides the usage of a selected enzyme reputation collection surrounding the meeting oligonucleotide collection. This oligonucleotide layout technique addresses the oligonucleotide awareness and pool complexity problems related to array-derived oligonucleotides at the same time as requiring no specialised chips or array synthesizers.