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URL: http://www.addgene.org/55218
Proper Citation: RRID:Addgene_55218
Bacterial Resistance: Ampicillin
Defining Citation: PMID:28668116
Vector Backbone Description: Vector Backbone:pFastBac; Vector Types:Insect Expression; Bacterial Resistance:Ampicillin
Comments: In order to increase the efficiency of subcloning with the Series-11 Macrobac plasmids we developed a new strategy the uses LIC (Ligation Independent Cloning). The problem with the Series-11 ligation mediated subcloning was that when the plasmid size approached or exceeded 20 kb we had to screen many colonies to find a positive. We developed a strategy to use LIC for subcloning. Because this method uses no ligase, the cloning background associated with re-ligation of the empty plasmid are eliminated. 438-A is an untagged pFastBac LIC Subcloning vector. The 438 vectors use the LICvBac Forward primer and the LICv1 Reverse primer. LICvBacF - 5'-TACTTCCAATCCAATCG-3' (Note: ATG must be added to ORF) LICv1 Reverse - 5'-TTATCCACTTCCAATGTTATTA-3' As the target plasmid size gets >20kb you may have to increase the amount of DNA you anneal and/or transform. We have found this method gives no background colonies at all and 100% of the colonies we pick are positive. The cells we transform into are XL1Blues with a competency ~6x10^7cfu/ul. For more information, please see our website: http://qb3.berkeley.edu/qb3/macrolab/
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No rating or validation information has been found for pFastBac cloning vector with BioBrick PolyPromoter LIC Subcloning (438-A).
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Source: Addgene
