Guidelines for chimeric coat color estimation

ES cell lines are derived from different mouse strains therefore, chimeras generated by blastocysts injections will have different coat colors.

The ES cell lines derived from 129/Ola mouse strain are named E14, E14.1, E14tg2a, CGR8.8. Genotype of 129/Ola mouse strain is cch/cch, p/p, Aw/Aw (agouti chinchilla coat color). Injection of these ES cells lines into C57Bl/6 will generate tri-colored chimeras, which should be judged according to the following guidelines:

Grade A chimera coat color comprises of significant percentage of agouti (about 70%) with patches of chinchilla coat color. Grade A chimeras can exhibit about15% of chinchilla coat or be completely chinchilla. Variations of chinchilla can include slate or light gray coat color. Small percentage of black coat color contributed by embryo donor may be present (about 20%). Grade A chimeras can have pink eyes.

Grade B chimeras exhibit mostly agouti coat color with some patches of chinchilla (1%-15 %). Black coat color (contributed by embryo donor) should comprise no more than 30%. Some grade B chimeras can be 100% agouti.

Grade C chimeras exhibit mostly black coat color with patches of agouti coat color. Percentage of agouti coat color varies from 5% to 70%.

E14 Chimera Grade A

The ES cell line R1, derived from cross of 129/Sv x 129/Sv-Cp mouse strains and other ES cell lines such as D3, W4, AB2.2, AK7.1 derived from different substrains of 129 mouse strain have the agouti coat color.

Injection of these ES lines into C57Bl/6 blastocysts will generate chimeras, which should be judged according to the following guidelines:

Grade A chimeras exhibit high percentage of agouti coat color 70% to 100% agouti. Agouti color indicates percentage of ES cell’s contribution to the individual animal. Percentage of black color coat derived from embryo donor should not exceed 30%.

Grade B chimeras exhibit high percentage of agouti coat color 50%-70%. Percentage of black color coat contributed by embryo donor should not exceed 50%.

Grade C chimeras exhibit mostly black color coat with some percentage of agouti 1%-40%.

R1 Chimeras Grades A, B, C

Bruce 4 ES cell line was derived from C57Bl/6 mouse strain. Genotype of this line is C/C, P/P, a/a. Being injected into albino host embryos such as BALB/c or albino B6 this cell line will generate chimeras exhibiting patches of black coat color or shades of brownish-gray coat color when injected into FVB/N host embryos indicating percentage of ES cells contribution. These chimeras should be judged according to the following guidelines:

Grade A chimeras exhibit some white coat color with high percentage of black 50% to completely 100% black coat color (brownish/grey when FVB/N donor used).

Grade B chimeras exhibit mostly white coat color with some patches of black 30%-40% (brownish/grey when FVB/N donors used).

Grade C chimeras exhibit mostly white coat color with small percentage of black 1%-20% (brownish/grey when FVB/N donor used).

We recommend breeding of all available chimeras keeping in mind that sometimes it takes 4-5 litters of mice before germline transmission will occur.

Bruce 4 Chimeras Grades A & C

C2J4 ES cell line was created in Rockefeller University and temporarily offered to investigators on the experimental basis. C2J4 line derived from C57BL/6J-Tyrc-2J/J mouse line, also known as albino B6 mice. Being injected into C57Bl/6 blasocysts it generates chimeras exhibiting white patches of coat color indicating ES cell contribution to the genotype. Those chimeras should be judged according following guidelines:

Grade A chimeras exhibit some black coat color with high percentage of white 50% to completely 100% white coat color.

Grade B chimeras exhibit mostly black coat color with some patches of white 30%-40%.

Grade C chimeras exhibit mostly black coat color with small percentage of white 1%-20%.

C2J4 Chimera Grade B