4th chromosome balancers

As explained in An Introduction to Balancers, X, second and third chromosomes must have both multiple inversions and recessive lethal or sterile mutations to work effectively as balancers. Since meiotic crossovers normally do not occur between fourth chromosomes, inversions are not necessary for a fourth chromosome to act as a balancer. Any fourth chromosome with a recessive lethal or sterile mutation can serve as a balancer for another fourth chromosome.

The recessive lethal, fourth chromosome mutations that work best for balancing other mutations are those that are also associated with easily scored phenotypes. Here we have listed a few convenient classes of recessive lethal mutations.

ci[D] and ey[D] have dominant wing vein and eye phenotypes, respectively. They are the two mutations used most often for balancing other fourth chromosome mutations. Their phenotypes can be easy to score in some stocks, but they tend to get suppressed upon long-term culture. It is often necessary to outcross and rebuild stocks to recover strong ci[D] and ey[D] phenotypes. You can find a full list of ci[D] and ey[D] stocks by typing the allele names into the search on our home page.

ci[Ce-2] gives ocellar and ocellar bristle phenotypes, but, it has not been used much for making stocks.


T(1;4)B[S] is associated with a severe Bar-eyed (B[S])  phenotype and can be used in crosses to follow the inheritance of fourth chromosomes. It is not, however, recessive lethal or sterile, so it cannot be used for making stable stocks. It’s not particularly useful as a balancer, but it’s convenient in some crossing schemes.

sv[spa-Cat], bt[D] and Scn[1] are dominant, but their phenotypes usually have weak penetrance and expressivity, so they are less useful than other mutations.


Mutations in RpS3A or deletions removing it are associated with moderate-to-strong, dominant Minute phenotypes, but heterozygotes have erratic viability and sterility.

P{ActGFP}unc-13[GJ] is a ubiquitously expressed GFP marker that is nice for making stocks where balancer-bearing flies need to be identified in preadult stages. The construct also carries miniwhite, which allows balancers to be followed in w[-] backgrounds.


TI{TI}Crk[dsRed] expresses dsRed under the control of the 3xP3 regulatory sequence to give eye and ocellar fluorescence. The transgene insertion replaced Crk coding sequence to generate a recessive lethal mutation.


All Mi{ET1} insertions carry the 3XP3-GFP marker, which results in green eyes and ocelli. We have one insertion on a lethal chromosome


The attP target insertion M{3xP3-RFP.attP}ZH-102D expresses RFP in eyes and ocelli and phiC31-mediated insertions into it also show fluorescence. It is homozygous viable and fertile, so it is useful for following fourth chromosomes in crosses, but not for creating stable, balanced stocks.

TI{GMR-HMS04515}Gat[eya] expresses dsRNA for RNAi of eya under the control of GMR regulatory sequences to produce a strong eye morphology phenotype similar to those of dominant Dr mutations. The transgene insertion disrupts Gat to cause recessive lethality.

Lethal or sterile w[+]- or y[+]-marked transposon insertions can be useful for balancing mutations in w[-] or y[-] mutant backgrounds. To find stocks with the insertions below - 1) go to our Advanced Symbol Search, 2) type the construct name from below into the "Symbol contains" box, 3) change the "chromosome" dropdown to "4" (for Chromosome 4) and 5) click submit.

For w[+]-marked insertions, consider PBac{5HPw[+]}P{GSV1}P{GawB} and P{lacW} stocks. Some fourth chromosome deletions and homologous replacement alleles are also associated with miniwhite insertions.

For y[+]-marked insertions, consider PBac{3HPy[+]}Mi{MIC} and PBac{HpaI-GFP.A}  insertions.

For w[+]- and y[+]-marked insertions, consider P{SUPor-P} and P{EPgy2} insertions.

For w[+]- and GFP-marked insertions, consider P{Act-GFP}unc-13[GJ] (see Mutations associated with GFP expression above).

Flies with three fourth chromosomes are viable, fertile, healthy and visibly indistinguishable from flies with the usual two chromosomes. Consequently, diplo-4 gametes are not uncommon in crosses and can lead one to misinterpret complementation results.

Likewise, flies with a single fourth chromosome are viable. Because the fourth chromosome carries the Minute gene RpS3A, a fly with a single fourth chromosome will show Minute phenotypes. They will have macrochaetae one-half to two-thirds the normal length and thinner than usual, their bodies will be slightly smaller than usual, and they will be less viable and fertile. Haplo-4 progeny typically appear at a rate of ~1 in 5,000 due to spontaneous meiotic nondisjunction.

Related links  Balancers   All Blue Balancers