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cutB

◈ Southern blotting (probe : ORF2L PCR product). probe(970 bp). orf6. orf2. orf5. orf4. cutB. cutC. cutA. Pst I (2290 bp). Nco I (5223 bp). Nco I 와 Cla I (4701 bp). probe(970 bp). orf6. orf2. orf5. orf4. cutB. cutC. cutA. Pst I (2290 bp). Nco I (5223 bp). Cla I (5121 bp).

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cutB

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  1. ◈ Southern blotting (probe : ORF2L PCR product) probe(970 bp) orf6 orf2 orf5 orf4 cutB cutC cutA PstI (2290 bp) NcoI (5223 bp)

  2. NcoI와 ClaI (4701 bp) probe(970 bp) orf6 orf2 orf5 orf4 cutB cutC cutA PstI (2290 bp) NcoI (5223 bp) ClaI (5121 bp)

  3. cutC cutA cutB orf2 cutR orf1 orf4 orf5 orf6 1kb 2290 bp PstI deleted orf2 partial cutA partial orf4 pMSM2 kanR hygR sacB Single crossover : selection on hygromycin containing media partial orf4 deleted orf2 partial cutA sacB cutA hygR orf4 orf5 orf2 cutB cutC kanR partial orf4 deleted orf2 1828 bp 1930 bp sacB cutA hygR cutB cutC partial cutA orf5 orf2 orf4 kanR Double crossover : selection on sucrose containing media deleted orf2 cutA cutB cutC Molecular Microbiology Lab., Yonsei University 1828 bp orf5 orf4

  4. cutC cutA cutB orf2 cutR orf1 orf4 orf5 orf6 1kb 5223 bp NcoI deleted orf2 partial cutA partial orf4 pMSM2 kanR hygR sacB Single crossover : selection on hygromycin containing media partial orf4 deleted orf2 partial cutA sacB cutA hygR orf4 orf5 orf2 cutB cutC kanR partial orf4 deleted orf2 15524 bp sacB cutA hygR cutB cutC partial cutA orf5 orf2 orf4 kanR Double crossover : selection on sucrose containing media deleted orf2 cutA cutB cutC Molecular Microbiology Lab., Yonsei University orf5 orf4 4764 bp

  5. ◈ Southern blotting (probe : ORF2L PCR product) 1 2 3 4 5 6 1 2 3 4 5 6 #1 : 1 kb ladder #2 : M. smegmatis wild type / PstI (2290 bp) #3 : M. smegmatisorf2 deletion mutant-1 / PstI (1828 bp) #4 : M. smegmatisorf2 deletion mutant -2/ PstI (1828 bp) #5 : M. smegmatisorf2 deletion mutant -10/ PstI (1828 bp) #6 : 1 kb ladder

  6. ◈ Southern blotting (probe : ORF2L PCR product) 1 2 3 4 5 6 1 2 3 4 5 6 #1 : 1 kb ladder #2 : M. smegmatis wild type / NcoI (5223 bp) #3 : M. smegmatisorf2 deletion mutant-1 / NcoI (4764 bp) #4 : M. smegmatisorf2 deletion mutant -2/ NcoI (4764 bp) #5 : M. smegmatis orf2 deletion mutant-10 / NcoI (4764 bp) #6 : 1 kb ladder

  7. ◈ Southern blotting (probe : ORF2L PCR product) 1 2 3 1 2 3 #1 : 1 kb ladder #2 : M. smegmatis wild type / PstI (2290 bp) #3 : M. smegmatisorf2 deletion mutant / PstI (1828 bp)

  8. ◈ Southern blotting (probe : ORF2L PCR product) 1 2 3 4 5 6 1 2 3 4 5 6 #1 : 1 kb ladder #2 : M. smegmatis wild type / PstI (2290 bp) #3 : M. smegmatisorf2 deletion mutant / PstI (1828 bp) #4 : M. smegmatis wild type/ NcoI (5223 bp) #5 : M. smegmatisorf2 deletion mutant/ NcoI (4764 bp) #6 : 1 kb ladder

  9. ◈M. smegmatisorf2¯ deletion mutant : Southern blotting ◈ construction of vectors for the other M. smegmatisorf¯deletion mutants ☞ orf1, orf5, orf6, Rv3676 andcutR ◈ Northern blotting and RT-PCR ◈ comparison of the growth curve between M. smegmatiswild type and M. smegmatisdeletion mutants (orf2¯ , orf4¯ ) in SMB + 30% CO

  10. ◈ construction of vectors for the other M. smegmatisorf¯deletion mutants orf1 orf5 orf6 cutR, Rv3676

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