Edinburgh/30 June 2008

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Revision as of 14:26, 20 August 2008

Edinburgh iGEM 2008

Overview Step1 Step2

Week 3

Monday 30 June 08

Plasmid DNA minipreps:
- M1 to M6: pSB1A2+dxs white colonies from plate 11
- M7: BABEL1+glgC white colony from plate 12
- M8 to M12: BABEL2+glgC white and pale blue colonies from plate 13
M1 to M6 were digested with EcoRI/PstI (Gel 2). The expected pattern is pSB1A2 vector band at 2.04kb (2079 bp less prefix and suffix) and dxs insert band around 1.9kb. M1 showed bands around 1.1kb and 2.1kb. M5 showed no DNA. The other four showed a vector-like band around 2kb and a fainter, fuzzier band around 3kb, possibly a 'ghost' band. Thus none of the clones show the expected pattern of bands (although it is conceivable, since the vector and insert bands are so close in size, that they may be lying on top of each other; dxs has an internal EcoRV site at +504 and two HindIII sites at +606 and +1230, whereas pSB1A2 lacks such sites so this could be used to check). In conjunction with the total lack of growth on the appY plates, this suggests that something went wrong in the cloning procedure. The next step is to run the remaining ligation material on a gel and see what it looks like.

@@ Line 292: / Line 292: @@
 ==== Monday 30 June 08 ====
-* Plasmid DNA minipreps:<br />
+* Plasmid DNA minipreps:
-'''M1 to M6:''' pSB1A2+''dxs'' white colonies<br />
+** '''M1 to M6''': pSB1A2+''dxs'' white colonies from plate 11
-'''M7:''' Babel1+''glgC'' white colony<br />
+** '''M7''': BABEL1+''glgC'' white colony from plate 12
-'''M8 to M12:''' Babel2+''glgC'' white and pale blue colonies<br />
+** '''M8 to M12''': BABEL2+''glgC'' white and pale blue colonies from plate 13
-Minipreps M1 to M6 were digested with EcoRI and PstI ('''Gel 2'''). The expected pattern is pSB1A2 vector band at 2.04kb (2079 bp less prefix and suffix) and ''dxs'' insert band around 1.9kb. M1 showed bands around 1.1kb and 2.1kb. M5 showed no DNA. The other four showed a vector-like band around 2kb and a fainter, fuzzier band around 3kb, possibly a 'ghost' band. Thus none of the clones show the expected pattern of bands (although it is conceivable, since the vector and insert bands are so close in size, that they may be lying on top of each other; ''dxs'' has an internal EcoRV site at +504 and two HindIII sites at +606 and +1230, whereas pSB1A2 lacks such sites so this could be used to check). In conjunction with the total lack of growth on the ''appY'' plates, this suggests that something went wrong in the cloning procedure. The next step is to run the remaining ligation material on a gel and see what it looks like.
+* M1 to M6 were digested with EcoRI/PstI ('''Gel 2'''). The expected pattern is pSB1A2 vector band at 2.04kb (2079 bp less prefix and suffix) and ''dxs'' insert band around 1.9kb. M1 showed bands around 1.1kb and 2.1kb. M5 showed no DNA. The other four showed a vector-like band around 2kb and a fainter, fuzzier band around 3kb, possibly a 'ghost' band. Thus none of the clones show the expected pattern of bands (although it is conceivable, since the vector and insert bands are so close in size, that they may be lying on top of each other; ''dxs'' has an internal EcoRV site at +504 and two HindIII sites at +606 and +1230, whereas pSB1A2 lacks such sites so this could be used to check). In conjunction with the total lack of growth on the ''appY'' plates, this suggests that something went wrong in the cloning procedure. The next step is to run the remaining ligation material on a gel and see what it looks like.