Pipeline Status Explanations

The following list defines the different statuses that we report on the constructs in our pipeline.

Mice - Genotype confirmed (M-GC)

The genotype of testcross offspring has been confirmed molecularly.

Mice - Germline transmission (M-GLT)

Test crosses of chimaeras indicate germline transmission of the ES cell mutation based on coat color.

Mice - Microinjection in progress (M-MIP)

Targeted ES cells have been microinjected for the purpose of creating mutant mice for distribution.

ES Cells - Targeting Confirmed (ES-TC)

Correctly targeted ES cell clones have passed LR-PCR/sequenced-based QC. These ES cell clones can be requested from the EUCOMM repositories.

ES Cells - Targeting Unsuccessful - Project Terminated (ES-PT)

Electroporation of the final targeting construct containing a promoter-driven selectable cassette yielded an insufficient number of colonies for analysis. Work on this particular gene will cease and the gene may be passed to Regeneron or Norcomm.

Vector Complete - Project Terminated (TVC-PT)

A project which has targeting vector but after re-sythesis still no successful es cell, so it terminated.

ES Cells - No QC Positives (ES-NQP)

ES Cells - Electroporation Unsuccessful (ES-EU)

ES cell clones were picked and screened, but did not pass LR-PCR/sequenced-based QC. These ES cell clones may be re-screened with additional genotyping primers.

ES Cells - Electroporation in Progress (ES-EP)

Electroporation of the final targeting construct is underway.

Vector - DNA Not Suitable for Electroporation (ES-DNA)

A final targeting vector (post Gateway) has been constructed, has passed sequence-based QC, but has failed gel-based QC prior to electroporation (low DNA yield, incomplete digestion or multiple DNA bands). These vectors will be re-prepped for electroporation and can be requested from the EUCOMM repositories.

Vector Complete (TVC)

A final targeting vector (post Gateway) has been constructed, has passed sequence-based QC, and is available for use in ES cells. These vectors can be requested from the EUCOMM repositories.

Vector - Initial Attempt Unsuccessful (TV-IAU)

The construction of a final targeting construct, following the Gateway exchange reaction, was not successful or did not pass sequence-based QC; pending a second attempt.

Vector Unsuccessful - Alternate Design in Progress (TV-ADP)

The construction of the targeting vector was not successful or did not pass sequence-based QC. Recombineering will be repeated using an alternative valid conditional design (allele).

Vector Unsuccessful - Project Terminated (TV-PT)

A project is terminated when two attempts at producing a valid conditional design (allele) have failed.

Vector Construction in Progress (TVIP)

Oligos for recombineering have been ordered for the construction of the targeting vectors.

Design Completed (DC)

The gene has been manually annotated by the Havana group and the design of a conditional allele has been generated.

Design Not Possible (DNP)

The gene has been manually annotated by the Havana group but the design of a conditional allele has not been possible. Reasons for this may include the absence of frame-shifting (critical) exons, the introns flanking critical exons are too small, too few exons (our vector design strategy requires the presence of at least two exons), etc. Requests are usually put on hold in this status unless other designs (e.g. deletion alleles) are specified.

VEGA Annotation Requested (AR)

The first step in the process of making an allele is to manually annotate the gene. This is carried out by the Havana group and the annotation is made publicly available on the Vega website at the next release.

Alternate Design Requested (ADR)

The construction of the Gateway-adapted intermediate vector was not successful or did not pass sequence-based QC. Recombineering will be repeated using an alternative valid conditional design.

Design Requested (DR)

The gene has been manually annotated by the Havana group but has yet to enter the design process.

Withdrawn From Pipeline (W)

The EUCOMM project(s) will not, or has ceased to, work on this gene. This may occur for a number of reasons for example if it is registered as having 6 or more hits in the TIGM database.

Transferred to NorCOMM (TN)

Work on this gene has been transferred to the Norcomm project. This most often occurs when the Norcomm pipeline receives a request to prioritise this gene.

Transferred to KOMP (TK)

Work on this gene has been transferred to the KOMP project because the design of a conditional allele (vector) has not been possible. This can occur for a number of reasons including the absence of frame-shifting (critical) exons; the introns flanking the critical exons (normally 1 and 2) are too small or there are too few exons (our vector design strategy requires the presence of at least two exons).

On Hold (H)

Genes with this status are on the list of genes to be targeted by the Eucomm pipeline but they have yet to receive any attention. It is the default status when a prioritisation request is entered into the database.

ES Cells - Conditional Gene Trap (ESC-GT)

DNA sequence from the gene trap clones is used to identify the intronic insertion site, sequence from the targeting vectors is used to define the precise allele structures and BAC ends are used to position the BACs in the genome. This information is mapped onto the mouse genome sequence using standard methods.

Regeneron (REG)