Ac-bti95004 Southern blot verification

Ac-bti95004 Southern blot verification:
Lane 1 = TB95-4.2
Lane 2 = TB95-4.3
Lane 3 = P-vv
Lane 4 = W22
Lane D = DIG VII ladder

Restriction Enzyme: EcoR1
Expected band size:. ~3.8 kb

Southern verification notes: Genomic DNA isolated from lines carrying Ac-bti95004 (lanes 1 & 2) or the progenitor lines (lanes 3 & 4), was digested with EcoR1, fractionated on agarose gels and transferred to nylon membranes. A DIG-labeled flanking sequence probe of 180 bp was generated using the Ac-end primer (Ac9003'end: 5'-CGTTACCGACCGTTTTCATCCCTA-3') and flanking sequence primer (p31M: 5'-GATTCCACCCCAGCTCTGAAAGTA-3') . Because EcoR1 cuts within the Ac element, the EcoR1 fragment is shifted in the lines containing Ac-bti95004 relative to the progenitor lines. Detailed descriptions of the Southern blot verification can be found in Brutnell and Conrad, 2003, Methods in Molecular Biology 236:157-76.


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