Editing Yijia LI 2 19 2018

Date:2/20/2018

{| border="1" cellspacing="1" cellpadding="1" style="width:500px;"
|-
| #
| Template
| 2X PCR Mix
| Primers
|-
| 3027
| 2492-1,2,3
| Q 5&nbsp;
| 3020-Xba1-Nhe1-F + 3027-2b-Not1-Age1-R<br/>
|-
| 3076
| 3003 H-4,5,6
| Q 5
| 3076-myc-BsrG1-F&nbsp;+ 3076-myc-Nhe1-Kpn1-R<br/>
|}

[[File:PCR of 2492 3003 H.JPG|RTENOTITLE]]



Date:2/21/2018

{| border="1" cellspacing="1" cellpadding="1" style="width:500px;"
|-
| #
| Template
| style="width: 156px;" | Restriction Enzymes
| Note
|-
| 1-5
| 3154 pMiniT2
| style="width: 156px;" | EcoR I
| wrong enzymes
|-
| 7,8,10
| 3021 pMiniT2
| style="width: 156px;" | EcoR I
| <br/>
|-
| 11,14
| 3027 pMiniT2
| style="width: 156px;" | EcoR I
| <br/>
|-
| Aim:
| Test if we succeed to put insert into pMiniT 2
| style="width: 156px;" | <br/>
| <br/>
|}

Digestion of 3154, 3021 or 3027 pMiniT2



[[File:Digestion of 3154 3021or 3027.JPG|RTENOTITLE]]



Date:2/22/2018

{| border="1" cellspacing="1" cellpadding="1" style="width:500px;"
|-
| #
| products
| Restriction Enzymes
| Note
|-
| 1,2
| 3010 L pMiniT2
| EcoR I
| 1
|-
| 3,4,5
| 3001 pMiniT2
| EcoR I
| 4
|-
| 6,7,8,9,10
| 3076 pMiniT2
| EcoR I
| 7
|-
| 11,12
| 3027 pMiniT2
| EcoR I
| 11
|-
| Destrisprtion
| We did miniprep and digested these products to test if we were sucessful.
| 1,4,7,11 to do sequencing
| <br/>
|}

[[File:Digestion of 3010 L 3001 3027 3076.JPG|RTENOTITLE]]



Date:2/23/2018

Digestion Test for 3154 Restriction enzymes:BamH I+ Kpn I

[[File:DigestionTest for 3130.JPG|RTENOTITLE]]



Digestion for 3154 from miniprep

Restriction Enzyme:Kpn I+ BamH I

[[File:Digestion for 3154.JPG|RTENOTITLE]]

Result: good, ordered for sequencing;sequencing result is good too.

Digestion of Vector 3154 for 3129 ligation Cut by Kpn I + Not I

[[File:Digestion of Vector 3154 for 3129 .JPG|RTENOTITLE]]

We did ligation, but seems like the colonies are too much in a whole plate, so we planed to repeat but still picked up three colonies in this bad plate.

Digestion of Vetcor and Insert for 3129 ligation

[[File:Digestion of 3129 for ligation.JPG|RTENOTITLE]]

Digestion of Vector and Insert for 3027 Ligation

Restriction Enzymes: Nhe I+ Not I

[[File:Digestion of 3027 for ligation.JPG|RTENOTITLE]]

Because it looks like that the band of insert is not desired, so we repeated the digestion.

Another Digestion of Vector and Insert for 3027 Ligation

Restriction Enzymes:Nhe I+ Not I

[[File:Digestion of 3027 Ligation.JPG|RTENOTITLE]]

We got the similar band of the old one.So just cut the very light color band out,not sure, so cut both two light bands.

Digestion of insert from 3076 pMiniT2&nbsp;&nbsp;for 3076 ligation

Restriction Enzymes:BsrG I+ Kpn I

[[File:Digestion of Insert 3076- pMiniT2 for 3076.JPG|RTENOTITLE]]

Not sure about the band, So planned to repeat next week.

PCR of 2493 to get the insert for 3021

2X PCR Mix:Platinum HiFi

Primers:3021-F2&nbsp; + 3021-Age1-Xba1-R , Vector: BssHII - Nhe1

[[File:PCR of insert 2493 for 3021.JPG|RTENOTITLE]]

Good , so cut them and put into tube and add QG buffer
[[Category:My ELN]]