Strains: Escherichia coli (plasmids all in DH10B)
All as stabs in LB
+ 40 mg/L kanamycin (BIBAC2, pCH20)
+ 10 mg/L tetracycline (pCH30, pCH32)
Strains: Agrobacterium tumefaciens
|COR314||UIA143 pMP90 pCH32 BIBAC2|
|COR356||UIA143 pMP90 pCH32 BIBAC2.H150|
COR366 LBA4404 pCH32 BIBAC2
stab, LB + 50 mg/L kanamycin + 2 mg/L tetracycline
COR308 UIA143 pMP90 pCH32
stab, LB + 5 mg/L tetracycline
Biological Materials Transfer Agreement _____
Maps: BIBAC2, pCH20, pCH30, pCH32 _____
Liquid cultures of A. tumefaciens strains, should have antibiotics added at the Concentrations indicated for the stabs. In particular, pCH30 and pCH32 are not stable in the absence of selection (tetracycline).
To select for inserts in the sacB gene we plate out E. coli cells on LB + 40 mg/L kanamycin + 5% sucrose. [For library preparation we use LB + 80 mg/L kanamycin, to reduce a common contaminant in commercially available DH10B ElectroMAX competent cells (BRL).]
The sacB gene also works in Agrobacterium. So Agrobacterium strains that contain the BIBAC vector without insert will be sensitive to high levels of sucrose. Check your co-cultivation medium; if it contains high levels of sucrose, then the Agrobacterium cells will be killed. We substituted glucose with no problems. This is not a problem for BIBAC + insert, since the sacB gene has been inactivated.
You can use your standard plant transformation protocol. The only modification that we made (mentioned in the PNAS paper) was to increase the concentration of the Agrobacterium cells/ml that was used to dip the tobacco leaf strips. This did improve efficiency. We are currently testing this modification for tomato transformation.
BIBAC technology and related materials are available for licensing. The "intellectual property" that is the basis of BIBAC technology is the "backbone" of the binary-BAC plasmid. The "backbone" plasmid (pCH20) does not contain any plant selectable markers. Cornell expects that (most) potential licensees will want to introduce proprietary markers. The BIBAC vector was designed with this in mind, and can be easily modified to specifications.
For non-profit organizations to request BIBAC materials for research purpose only, please contact the Center for Technology Licensing at Cornell University (CTL) at firstname.lastname@example.org under the subject of D1639 material request. A Material Transfer Agreement will be prepared according the request. Please be advised that due to the fact that the inventor of BIBAC is no longer at Cornell, a fee of $240 will be charged to reimburse the expense in maintaining the materials and in shipping and handling. For commercial entities to request BIBAC materials and license information, please contact Carolyn Theodore (email@example.com) at CTL.
Please contact either of these individuals at Syngenta to obtain the Mogen strains:
Once you have permission from Syngenta to use MOG101 (this strain was formerly owned by MOGEN International), then all of the strains derived from MOG101 by Carol Hamilton will be available for your use.
MOG101 derived materials
Derivatives of MOG101 [C58 pMOG101]
UIA143: recA- deletion of C58
|UIA143 pMOG101 pCH*||LB + Tet5|
|UIA143 pMOG101 pCH* BIBAC*||LB + Kan50 + Tet5|
recA + (MOG101) derivatives available by request