3009 pCMV5-Flag-H-IgG2b

Date: 1/24/2018

Digest miniprep DNA with Xho1

vector: pMiniT2

1/25/2018

Miniprep DNA in pMiniT2 vector digested with Xho1

Digest of 3009pMiniT

Enzymes:BsrGI+NheI

Time:1hour

Buffer:NEB2

Digest of 3009 pMiniT

Enzymes:XhoI+NheI

Time:30min

Buffer:Fastfigest

Subclone of 945

1:1727 XbaI+NcoI 173  Cutsmart

2:1834 XbaI+NcoI 173  Cutsmart

3:945pMiniT XbaI+NcoI 1326  Cutsmart

4:3009pMiniT BsrGI+NheI 371   NEB 2

Time:30min

Result:1727 can see band as vector, 945pminiT2 can see band as insert, cut out and combine,low concentration of 309pMiniT

[[File:Subclone of 945.JPG|RTENOTITLE]

Date:3/15/2018

3009pMiniT2 digestion test 1

Restriction enzymes:BsrG I+NheI

Buffer:NEB 2

Time:30min

Result:seems Lane 4 has band

3009pMIniT2 digestion test 2

Restriction enzymes:XhoI

Buffer:Fastdigest

Time:10min

7:3009-4pminiT2 (sequenced)

Result:some of them have bands, Lane 4 has band too.

3009pMiniT2 digestion Test 3

1 3009-4 BsrGI +KpnI

2 3009#5 BsrGI +KpnI

3 3009#5 BsrGI +NdeI

4 3009#5 NdeI +NheI

Buffer:NEB 2

Time:30min

Analysis of 3009-4 in pMiniT2

3009-4 in pMiniT2 digested with Nhe1-BsrG1, Kpn1-BsrG1

Date: 3/26/18, by Paul

Buffer: FastDigest

37 oC 1.5 hour

don't know if Nhe1 sitte is present, need to transform INV110