Abstract
Phage DNA isolation from genomic and cDNA libraries is time-consuming and cumbersome. Our work reports on the use of a pair of λ gt11 primers to amplify by PCR any insert DNA that is cloned into theEco RI restriction site. Using crude phage pools and direct phage plaques from a corn root cDNA library, we amplified and cloned DNA fragments. The amount of amplification products was similar when phage DNA and phage pools were used as templates. Amplified fragments could be purified, cloned and sequenced by a relatively simple procedure, thus allowing rapid detection and analysis of inserts in a large number of phage plaques.
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Abbreviations
- DIG:
-
digoxigenin
- LB:
-
Luria-Bertani medium
- PCR:
-
polymerase chain reaction
- PHBCS/H:
-
consensus probe for plant hemoglobin
- PPP:
-
primary phage pool
- SPP:
-
secondary phage pool
- SM:
-
SSC, seeMaterials and Method
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Arredondo-Peter, R., Bonic, A., Sarath, G. et al. Rapid PCR-based detection of inserts from cDNA libraries using phage pools or direct phage plaques and lambda primers. Plant Mol Biol Rep 13, 138–146 (1995). https://doi.org/10.1007/BF02668785
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DOI: https://doi.org/10.1007/BF02668785