Restriction enzymes
-------------------
-We'll use the following [restriction enzymes][renz]
+We'll use the following [restriction enzymes][renz]:
[BamHI][]
...................................
where the "other" sequence is the result of the BamHI/BglII splice.
-Expanding the "-SP-" abbreviation around the SP joint:
+Expanding the `-SP-` abbreviation around the SP joint:
....SP,other_.HindIII. SP.....
Leu Met Asp Leu Ser Phe Met Arg
This cDNA match generates an amino acid starting with LIKVEK instead
of the expected LIEVEK, but the LIKVEK version matches amino acids
12677-12765 in [Q8WZ42][] (canonical titin), and there is a natural
-variant listed for [12679 K->E][var].
+variant listed for [12679 K→E][var].
Interestingly, this sequence contains a PstI site at nucleotides 16220
through 16225. None of our other restriction enzymes have sites in
Since I'm stuck trying to get I27 into either plasmid, let's try and
work backward from
- Met-Arg-Gly-Ser-(His)₆-Gly-Ser-(I27-Arg-Ser)₇-I27-Cys-Cys
+ Met-Arg-Gly-Ser-(His)₆-Gly-Ser-(I27-Arg-Ser)₇-I27-...-Cys-Cys
#### <a id="BglII-BamHI-joint">BglII/BamHI joint</a>
The beginning of this sequence looks like the start of pQE30's linker,
so we'll assume the final plasmid was:
- Met-Arg-Gly-Ser-(His)₆-Gly-Ser-(I27-Arg-Ser)₇-I27-Arg-Ser-Cys-Cys
-
remote ... ,RGS-His epitope__________________. ,BamHI. I27...
... Met Arg Gly Ser His His His His His His Gly Ser Leu Ile ...
??? ... ATG AGA GGA TCG CAT CAC CAT CAC CAT CAC GGA TCC CTA ATA ...