Basics

Currently Mobix Lab orders oligos from Integrated DNA Technologies (IDT) Portal

Oligo synthesis proceeds in the 3' to 5' direction with the 3' base covalently attached to a solid support and utilizing cyanoethyl phosphoramidite nucleosides. As the coupling efficiency of each base addition step is less than 100% (normally between 98% and 99%) there is always a portion of short terminated oligos after synthesis.

Purification

Size

% full length (at 98% coupling efficiency)

Applications

Desalting

20mer
30mer
40mer
50mer
60mer
70mer
80mer
90mer
100mer

65
53
43
35
30
24
20
16
13

Sequencing, PCR

>50 desalt not recommended unless you intend to purify yourself

HPLC

10 – 100mer

Approx. 85%

Mutagenesis, cloning,     any other applications

 

Purification

Desalted oligos will contain the full-length oligo, as well as the smaller failed synthesis DNA fragments, which were capped due to failed incorporation of the next base.  These oligos are adequate for most applications, such as DNA sequencing, PCR amplification, etc. 

HPLC achieves a higher purity level (approximately 85%) but with a significant decrease in yield. 

PAGE purified oligos are the most pure (>90%) but yield is also quite low.

 

Modifications

For information about all DNA and RNA oligo modifications available from IDT, please visit:

https://www.idtdna.com/pages/products/custom-dna-rna/oligo-modifications

 

Degenerate Primers

Degenerate oligos are synthesized to contain equimolar mixtures of two or more different bases at a given position within the sequence. Mixed bases are often incorporated into oligonucleotide probes designed to hybridize to an unknown gene that encodes a known amino acid sequence. Since the genetic code is degenerate (e.g., Histidine could be encoded by CAC or CAT), the oligo probe is prepared with C or T at the degenerate site. Oligos with mixed bases are also useful in random mutagenesis and combinatorial chemistry. Degenerate oligos are produced for no additional charge (except for degenerate bases, where a hand-mixed variant is desired. Check the IUB codes for mixed bases.

 

IUB Codes

CodeDefinitionMnemonic
AAdenineA
CCytosineC
GGuanineG
TThymineT
RAGpuRine
YCTpYrimidine
KGTKeto
MACaMino
SGCStrong
WATWeak
BCGTNot A
DAGTNot C
HACTNot G
VACGNot T
NAGCTaNy

 

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