Crm at 23:17, 13 March 2018

2018-03-13T23:17:18Z

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== Date:3/5/2018 ==

3021:1-4 restriction enzymes:BssHII+XbaI

3027:5-8 restrcition enzymes:NotI+NheI

Buffer:Fastdigest

Time:16min

'''Result:'''seems good, but the result of sequencing is not,the product were actually are '''undigested vectors,3016pCEL0,3017pCEH0. so we collected them for other using.'''

[[File:Miniprep Digestion 3021 3027.JPG|RTENOTITLE]]

== Date:3/6/2018 ==

'''Subclone of 3027:'''

Vector:3017pCEH0

Insert:3027pMiniT2

restriction enzymes:MheI+NotI

Time:30 min

[[File:Subclone of 3027.JPG|RTENOTITLE]]

'''Result''':So we cut out the band and did subclone

'''Mini prep Digestion of 3162_3163_3164'''

3162:5-8 Not I+BamH I

3163:9-12 BsrG I+Not I

3164:13-16 BssH II+Nhe I

Buffer:Fastdigest

Time:30 Mins

'''Results:we selected 6,9,16 to do sequencing, all of the sequencing results are good and we labled the plasmid and did bacterial stock.'''

[[File:Miniprepdigest of 3162 3163 3164.JPG|RTENOTITLE]]

'''subclone test:'''

'''because miniprep digestion of 3162_3164_3165 has some good results, so we selected other sites to cut the DNA to doublecheck'''

3162:BssHII+KpnI 330

3163:NotI+KpnI 1101

3164:NheI+KpnI 377

[[File:Subclone test of 3162 3163 3164.JPG|RTENOTITLE]]

'''Test for 3017pCEH0 digestion:'''

'''because after digesting, three bands showed up in the gel.'''

Buffer:Fastfigest

Time:30min

Vector:3017pCEH0

'''Results:NheI may have another sites in the vector. becase NheI cutting has tha same result as NoyI+NheI cutting.'''

[[File:Vector3017 digest.JPG|RTENOTITLE]]

'''Subclone digestion of vector 3016pCEL0'''

Restriction enzymes:BssHII+NheI

Time:30min

Buffer:Fastdigest

Result: not good, did not see the small band.

[[File:Subclone digest of 3016pCEL0.JPG|RTENOTITLE]]

'''945 PCR'''

2X PCR Buffer:Platinum Supermix HiFi

Template:1281

Primer:945 F+ 945 R

'''Results:good, cut out and did gel purfication to prepare next step.'''

[[File:945PCR.JPG|RTENOTITLE]]

== Date:3/7/2018 ==

'''Miniprep digestion:'''

3021pMiniT2:1-4

Restriction enzymes:EcoRI

3161:5-8

Restriction enzymes:NdeI+XhoI

'''Results: '''we sent 3021pminiT2 to do sequencing

3161 is not good.

[[File:Digestion of 3161 3021pMiniT2.JPG|RTENOTITLE]]

'''PCR of 947'''

Lane1,2:Fragment 1 Primer:945 F+947 R

Lane3,4:Fragment 2 Primer:947 F+945 R

Buffer:Platinum HiFi supermix

'''Results:good, cut out and do next PCR'''

[[File:PCR of 947.JPG|RTENOTITLE]]

second PCR of 947

Primer: 945 F+945 R

Buffer:Patinum HiFi supermix

'''Results:good, cut out the first band '''

[[File:Second PCR of 947.JPG|RTENOTITLE]]

== Date:3/8/2018 ==

'''Miniprep digestion of 3021:'''

Buffer:Fastdigest

Time:15Min

Restriction enzymes: BssHII+NheI

'''Results:the second lane seems like a digested band'''

[[File:Miniprepof 3021.JPG|RTENOTITLE]]

'''Subclone of 3010;vector digestion'''

Vector:3130

Buffer: Fastdigest

Time:15min

'''Result:good cut out and combine with insert'''

[[File:Subclone of 3010.JPG|RTENOTITLE]]

== Date:3/9/2018 ==

'''subclone of 3132:'''

BsrGI+NheI

Buffer:Fastdigest

Time:15 min at 37oC

'''Result: good, cut out'''

[[File:Subclone of 3132.JPG|RTENOTITLE]]

'''subclone of 3168:'''

Restriction enzymes:BsrGI+NheI

Buffer:Fastdigest

Time:15 min at 37oC

'''Results: good,cut out and combine'''

[[File:Subclone of 3168.JPG|RTENOTITLE]]

'''subclone of 3152:'''

Restriction enzymes:BsrGI+NheI

Buffer:Fastdigest

Time:15 min at 37oC

'''Result: Vector is good, insert is not good, just cut out vector'''

[[File:Subclone of 3152.JPG|RTENOTITLE]]

'''subclone of 3167:'''

Restriction enzymes:BsrGI+NheI

Buffer:Fastdigest

Time:15 min at 37oC

'''Result: Vector is good, insert is not good, just cut out vector'''

[[File:Subclone of 3167.JPG|RTENOTITLE]]

'''subclone of 3131:'''

Restriction enzymes:BssHII+XhoI

Buffer:Fastdigest

Time:15 min at 37oC

'''Results: good,cut out and combine'''

[[File:Subclone of 3131.JPG|RTENOTITLE]]

'''subclone of 3009:'''

Restriction enzymes:BsrGi+KpnI

Buffer:Fastdigest

Time:1 hour at 37oC

'''Result:good cut out bands and combine'''

[[File:Subclone of 3009new.JPG|RTENOTITLE]]

'''subclone of 3135:'''

Restriction enzymes:BsrGi+KpnI

Buffer:Fastdigest

Time:1 hour at 37oC

'''Result:good cut out bands and combine'''

[[File:Subclone of 3135.JPG|RTENOTITLE]]

'''subclone of 3076:'''

Restriction enzymes:BsrGi+KpnI

Buffer:Fastdigest

Time:1 hour at 37oC

'''Result:vector is good cut out band, insert is not good.'''

[[File:Subclone of 3076.JPG|RTENOTITLE]]
[[Category:My ELN]]

Yijia LI 3 5 2018 - Revision history

Crm at 23:17, 13 March 2018