An Efficient DNA Sequencing Strategy Based on the Bacteriophage Mu in Vitro DNA Transposition Reaction

Table 1.

Sequencing the Mouse KCC2 Locus with the Mu System

Plasmid pSR11
Insert size (kb) 10.3
Vector size (kb) 2.9
Minitransposon used cat–Mu
E. coli cells used
 Strain DH5α
 Competence (cfu/μg pUC19 DNA) ∼107
Plasmid introduced into cells by electroporation
Fraction of standard reaction used in  electroporation 1:10
Number of ApR/CmRcolonies obtained 540
Number of clones screened by  restriction analysis 95 (100%)
  two plasmids apparent 2 (2%)
  one plasmid apparent 93 (98%)
Clones with one transposon 93 (98%)
 in vector 14 (15%)
 in insert 79 (83%)
Number of clones sequenced 71 (75%)
Total sequenced (bp) 68,143
Final sequence (bp) 10,288
Average read per sequence reaction
 (nucleotide) 494
Redundancy (fold) 6.6
  • Sequences retrieved using RP and UP primers are included.

  • These clones exhibited a restriction pattern consistent with cotransformation of two plasmids.

This Article

  1. Genome Res. 9: 308-315

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