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Balancer chromosomes are special, modified chromosomes used for genetically screening a population of organisms to select for heterozygotes. Balancer chromosomes can be used as a genetic tool to prevent crossing over (genetic recombination) between homologous chromosomes during meiosis. Balancers are most often used in Drosophila melanogaster (fruit fly) genetics to allow populations of flies carrying heterozygous mutations to be maintained without constantly screening for the mutations but can also be used in mice.^[1] Balancer chromosomes have three important properties: they suppress recombination with their homologs, carry dominant markers, and negatively affect reproductive fitness when carried homozygously.

History of balancer chromosomes

Balancer chromosomes were first used by Hermann Muller.^[2] In the modern usage of balancer chromosomes, random mutations are first caused, usually by feeding the larva ethyl methane sulphonate. When a phenotype of interest is observed, the line is crossed with another line containing balancer chromosomes to maintain their lineage.^[3] In one instance they were used to genetically screen a population of Caenorhabditis elegans. At this point in time scientists had already realized the benefits of being able to genetically screen populations of organisms for genetic study. Equally as important, they also realized that they could limit crossing over in these populations as well give them a very consistent genetic population.^[4]

The use of balancer chromosomes has evolved into a well known and widely used method for genetic screening of model organisms. They are even being used to investigate the role of heterochromatin packing and the effect it has on genes^[5] as well as studies on the effect telomeres have on gene silencing.^[6]

How balancer chromosomes work

To suppress crossing over, balancer chromosomes are the products of multiple, nested chromosomal inversions so that synapsis between homologous chromosomes is disrupted. This construct is called a crossover suppressor.^[7] If crossing over between a balancer chromosome and the balancer's homolog does occur during meiosis each chromatid ends up lacking some genes and carrying two copies of other genes. Recombination in inverted regions leads to dicentric or acentric chromosomes (chromosomes with two centromeres or no centromere). Progeny carrying chromosomes that are the products of recombination between balancer and normal chromosomes are not viable (they die).

Dominant markers such as genes for green fluorescent protein or enzymes that make pigments allow researchers to easily recognize flies that carry the balancer chromosome.^[8] By suppressing reproductive fitness when carried homozygously a balancer chromosome ensures that the population it is carried in does not become fixed for the balancer chromosome.

Balancer chromosomes always contain a lethal recessive allele. This means that if an organism receives two copies of the balancer chromosome, one from the mother and one from the father, then the organism will not live. So any organism that is homozygous for that chromosome will not live to pass on its genes. However, offspring that only get one copy of one balancer chromosome and one copy of a wild type or mutant chromosome will live to pass on its genes. After only a few generations the population will be entirely heterozygous so that you can be guaranteed of its genotype on at least those two chromosomes.

Balancer chromosomes also come with some sort of physical marker. This marker can be actually associated with the DNA in the chromosome such as the Green Fluorescent Protein that fluoresces in ultraviolet light, or it can be an easily distinguishable physical characteristic. These physical characteristics can be anything that is seen easily. In Drosophila melanogaster, for example, eye color and hair length are commonly used. This physical marker serves as a double check that you indeed have the heterozygous balancer chromosomes in the organism.^[9]

Naming convention

Balancer chromosomes are named for the chromosome they serve to stabilize and for the phenotypic or genetic marker the balancer carries.^[10] The naming of balancer chromosomes in D. melanogaster has been standardized as follows: the first letter of the chromosome's name represents the number of the chromosome it stabilizes. F stands for the first chromosome, S stands for second, and T stands for third (the small fourth chromosome does not undergo recombination and therefore does not require balancing.) This letter is then followed by an M, for "multiply inverted." The M is followed by a number to distinguish balancers of the same chromosome. Additionally, the genetic marker or markers of the balancer are listed after the name and separated by a comma. Generally mutations with easily observable dominant phenotypic traits that are often homozygous lethal are used to ensure that all progeny are heterozygous. For example, the commonly used "TM3, Sb" balancer is a balancer chromosome that stabilizes the third chromosome and carries a mutant Sb ("stubble") gene as a marker. All flies containing the TM3, Sb balancer will have shortened (or stubbly) hairs on the back of the fly, which are easily seen when viewed through a microscope. The 3 is to distinguish this from other third chromosome balancers, such as "TM1" and "TM2".

A line is said to be "double-balanced" if it is heterozygous for two different balancer chromosomes (for example, TM6,Tb/TM3,Ser) on a balancer chromosome and a homozygous lethal, heterozygous visible mutant on the other, wild-type chromosome (for example, D/TM3,Ser). Most balancer chromosomes also carry a recessive allele such as the "ebony" mutation that is only manifest in these stocks with two balancer chromosomes. These stocks are often used to provide sources of easily traceable traits when breeding two different lines together so that the correct progeny of each cross might be selected. Stocks double-balanced at both the second and third chromosomes in Drosophila are widely available.

Important scientific contributions using balancer chromosomes

Balancer chromosomes already give geneticists a reliable method for genetically screening organisms for a mutation and keeping that line constant. A new technique using balancer chromosomes is explored in the paper "The Autosomal Flp-Dfs Technique for Generating Germline Mosaics in Drosophila Melanogaster." This paper showed for the first time that it is possible to screen for a recessive mutation that only shows phenotype when homozygous. Using old balancer chromosome methods, genetic screening only allowed the picking out of heterozygous dominant mutations. This experiment uses clonal screening to detect homozygous individuals and keep them in a constant line.^[11]

They achieved this by using a gene isolated from yeast. This gene is called FLP recombinase and causes large chromosomal inversions. Through trial and error they found that the chromosomes could be recombined such that each had the recessive mutation while the other half contained half of a balancer chromosome with a physical marker and a lethal recessive. The other homolog did not contain the lethal recessive in the lines that survived. Figure one in the paper illustrates the screen. This new technique allowed recessive screening in 95% of the Drosophila genome. It also greatly improved yields in germ line mutations.^[11]

Another published paper that employed the use of balancer chromosomes is "Inhibition of RNA Interference and Modulation of Transposable Element Expression by Cell Death in Drosophila." This paper demonstrates the power of balancer chromosomes and what can be accomplished with genetically stable lines. A line was established that exhibited low levels of cell death and was named EGFPir hs-hid. The RNAi levels were analyzed and they found interesting results in the cells undergoing low levels of cell death and the surrounding cells in the tissue. They found that these cells would shut down their RNAi mechanism via maintaining RNA in a double stranded state. If RNA remains in a double stranded state then the RNAi mechanism of gene silencing is shut down.

The authors speculated that this response was an evolutionary trend toward redundant immune response against RNA viruses. If one cell is already undergoing cell death to attempt to stop spread of a virus, then the RNAi immune response has been ineffective. This causes another immune response that attempts to stop the virus which is binding double stranded RNA and keeping double stranded so that it cannot be transcribed into virus proteins. The mechanism of maintaining double stranded RNA is not known.^[12]

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