There have been few studies of spontaneous mutation accumulation in stocks under normal culturing practices. To undertake a longitudinal study of such sequence changes, we built stocks consisting of Y, second and fourth chromosomes isogenized from a variety of different wild type stocks in December 2022. In other words, the second and fourth chromosome pairs in each stock should have been completely identical at the time the stocks were established and a single Y chromosome should have been present in each stock. The X and third chromosomes were not isogenized and are likely highly heterozygous. We intend to analyze sequence changes over time by periodically sampling the stock populations.

We are making these stocks available to anyone who might want to use them.  

As shown in detail in the following crosses, we have extracted single second and fourth chromosomes from the ten “wild type” stocks listed above and homozygosed them using second chromosome balancers and dominant fourth chromosome markers. The Y chromosomes in the final stocks are also derived from these ten stocks. The X and third chromosomes in the final stocks are recombinant mixtures of chromosomal segments from the stocks used in the crosses. In other words, the Y, second and fourth chromosomes in the final stocks should have been isogenic in December 2022 while there was significant sequence diversity among the X and third chromosomes within a stock.

G0: 2555 CyO/sna[Sco] ♀♀ x 94597 w[1118]; TI{RFP[DsRed.3xP3.cUa]=FRT.Tub.GAL80.O}101F-DsRed+ ♂♂

 (This cross generated progeny for G1 Cross 2.)

G1 consisted of two parallel crosses:

G1 Cross 1: 7145 Df(2R)BSC39, cn[1] bw[1]/SM6a, bw[k1] ♀♀ x 90852 y[1] w[*]; TI{GMR-HMS04515}Gat[eya]/In(4)ci[D], ci[D] pan[ciD] ♂♂

G1 Cross 2: +/w[1118]; sna[Sco]/+; +/+; +/TI{RFP[DsRed.3xP3.cUa]=FRT.Tub.GAL80.O}101F-DsRed+ ♀ progeny of G0 cross x wild type ♂♂

(Both crosses generated progeny for the G2 cross. G1 Cross 2 was repeated for each of the ten wild type stocks.)

G2: +/y[1] w[*]; SM6a, bw[k1]/+; +/+; +/(TI{GMR-HMS04515}Gat[eya] or In(4)ci[D], ci[D] pan[ciD]) ♀ progeny of G1 Cross 1 x (w[+] or w[1118])/”Y”; sna[Sco]/”+”; +/”+”; TI{RFP[DsRed.3xP3.cUa]=FRT.Tub.GAL80.O}101F-DsRed+/”+” ♂ progeny of G1 Cross 2

(Multiple females were mated to a single male to set up a line. Ten lines were set up for each wild type strain using females carrying TI{GMR-HMS04515}Gat[eya] and ten were set up using females carrying the In(4)ci[D], ci[D] pan[ciD] chromosome. ”+” and “Y” indicate chromosomes from the wild type males in G1 Cross 2.)

G3: (y[1] or y[+]) (w[*] or w[+])/y[+] (w[1118] or w[+]); SM6a, bw[k1]/”+”; +/+; (TI{GMR-HMS04515}Gat[eya] or In(4)ci[D], ci[D] pan[ciD])/”+” ♀♀ x (y[1] or y[+]) (w[*] or w[+])/”Y”; SM6a, bw[k1]/”+”; +/+; (TI{GMR-HMS04515}Gat[eya] or In(4)ci[D], ci[D] pan[ciD])/”+” ♂♂

(A G3 cross was set up for progeny of each individual G2 male, i.e. each line presenting a single second chromosome and a single fourth chromosome from a progenitor wild type stock. )

A G4 cross was set up for each line to get a final stock without any second chromosome balancers or dominant fourth chromosome markers. y[1] and mutant w alleles may be segregating in each stock.

In the strain names, the middle letters represent the wild type progenitor: Hikone-A-W (HAW), Florida-9 (F9), Hikone-R (HR), Lausanne-S (LS), Samarkand (S), Urbana-S (US), DrosDel (DD, because the w[1118]-marked progenitor stock 5905 was generated by the DrosDel Project), Berlin-K (BK), Canton-S (CS, obtained from Stuart Newfeld) and Oregon-R-modENCODE (OR). The final letter in the code is the line identifier. The final letter in the strain name is the line identifier. Line names ending in A–J were generated using the TI{GMR-HMS04515}Gat[eya] dominant marker in the crosses above and lines ending in K–T were generated using the In(4)ci[D], ci[D] pan[ciD] chromosome.

All lines were maintained as populations of roughly four vials from January through July 2023. Since then, the thirty stocks above have been maintained by BDSC stockkeepers using routine practices: each copy of each stock is maintained as two vials, which are used alternately to establish new cultures every 14 to 18 days.