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Table 1 Comparison of screening BAC library by conventional PCR and by MT-PCR-HRM.

From: A simple, high throughput method to locate single copy sequences from Bacterial Artificial Chromosome (BAC) libraries using High Resolution Melt analysis

Compared features Screening by conventional PCR using 3D pooling strategy Screening by MT-PCR-HRM using 2D pooling strategy
Maximum number of 384-well plates to be pooled in one super pool 10 384-well plates 25 384-well plates
Number of super pools to be tested to identify a positive superpool (N: The total plate number of the BAC library) N/10 N/25
PCR reactions (PCR/PCR-HRM) needed to identify a positive super pool N/10 N/25
Reactions needed to identify the plate ID for one positive super pool 10 10
Reactions needed to identify the clone ID from one positive plate 40 18
Total number of reactions needed to get positive BAC clone ID from whole library N/10 + 50 N/25 + 28
Multiplexing possibility No Yes
Checking on agarose gel Needed NO
Cost 0.16 £/1 PCR reaction + cost for agarose gel electrophoresis 0.17 £/1 PCR-HRM reaction
Procedure duration to anchor 1 marker 12 h 3 h
Scoring data Manually from gel photos Semi-Automatic (figures and summary tables can be exported from HRM rotor system)