96-Well Confirmation Yeast PCR Protocol Protocol for 96-well confirmation Yeast PCR. Includes: Clonal purification; Generate a master plate (96-well format); Making a frozen backup stock; Confirmation PCR for one Row; ORF Specific Confirmation PCR --> "A-B" primers (upstream junction); Transfer template DNA to multiwell PCR plate; Prepare and dispense master mix for A-B PCR.
Analyzing Yeast Colonies by PCR Protocol Yeast colonies are suspended in complete PCR buffer and transferred to a thermal cycler for 35 cycles of PCR. The products of the amplification reaction are analyzed by gel electrophoresis.
Identification of End Clones in YAC Subclone Libraries Protocol To identify the YAC subclones containing both a human insert and a portion of either the left or right arm of the pYAC4 vector. Identification of these clones is necessary in order to do YAC chromosome walking, and is also useful in the determination of whether a particular YAC clone has a contiguous human insert or whether a co-cloning event has occurred. Vector arm sequences are identified using pBR322 fragments from a BamHI-PvuII double digest.
Oligonucleotide Details for PCR Information for oligonucleotide details required for PCR. Includes: Primer choice; UPTAG PRIMER; DNTAG PRIMER; UP_45 and DOWN_45 PRIMERS; UP_90 and DOWN_90 PRIMERS.