SapI Entry vectors

A series of over a dozen KanR vectors designed to simplify creating SapI Golden Gate entry plasmids for the assembly of targeting plasmids for C. elegans genome manipulation. The vectors are compatible with inserting sequences using many different approaches including traditional restriction enzyme cloning, Gibson assembly and BsaI Golden Gate Cloning (Figure 1). A detailed manual that describes the methods is available.

Figure 1. Alternatives for cloning inserts into Golden Gate ‘entry’ vectors.
Six alternative methods for creating Golden Gate entry vectors.  A) a BsaI Golden Gate approach, B) a SapI cloning approach, C) a Gibson assembly cloning approach, D)  a traditional double sticky-end cloning approach E) a multi-fragment BsaI Golden Gate approach, and F) a multi-fragment Gibson assembly cloning approach.  Restrictions sites: B= BsaI, R=EcoRI, H=HindIII, S= SapI.  Inserts are shown in red and the vector in black.

Below is a list of the available plasmids:

NM #NameSapI Slot (caps)
& BsaI overlap		.gb file
[all seqs]
3855DR274 sgRNA – BsaIgAAC-CAAc[seq]
3854DR274 U6p -BsaIgCAA-TGGc[seq]
3467DR274 5’ arm – BsaIgTGG-GCGa[seq]
3468DR274 CT – BsaIgGCG-ATGa[seq]
3469DR274 FP – BsaIgATG-AAGg[seq]
3707DR274 SEC- BsaIgAAG-GGTc[seq]
3456DR274 NT – BsaIgGGT-ACGc[seq]
3470DR274 3’ arm – BsaIgACG-GTAg[seq]
3967†pET28- FP -BsaIgATG-AAGg[seq]
3844DR274 sgRNA-U6p – BsaIgAAC-TGGc[seq]
3698DR274 5’ arm-CT – BsaIgTGG-ATGa[seq]
3701DR274 5′ arm- SEC -BsaIgTGG-GGTa[seq]
3773DR274 CT-NT – BsaI gGCG-ACGc[seq]
4617DR274 FP-SEC -BsaIgATG-GGTa[seq]
3869DR274 FP-NT – BsaIgATG-ACGc[seq]
3843DR274 SEC-NT – BsaIgAAG-ACGc[seq]
3702DR274 SEC-3’ arm – BsagAAG-GTAg[seq]
† low copy number vector for toxic inserts

Last updated 7-29-2023