Nuclear Primers

Brower and DeSalle (1994) have reviewed some useful nuclear primers. Bonacum et al. (2001) also report on primers for 12 nuclear genes. If primers were designed with T3/T7 sequences to streamline high throughput sequencing, both the linked and unlinked sequences are given. Below, we include information based on our recent publications.

26S proteostome regulatory subunit (26S) 

Zilversmit et al. (2002)

under construction

28S

This is a highly conserved ribosomal locus that has been used by Pelendakis and colleagues (1991; 1993) to infer higher-level phylogenetic relationships in Drosophilidae. This gene has also been used at species group (O'Grady 1999), subgenus (O'Grady and Kidwell 2002), and genus level ( Remsen and O'Grady 2002; Zilversmit et al. 2002) divergences in Drosophilidae. This gene evolves too slowly and shows too much homoplasy at sites that do change to be useful.

under construction

Acetylcholinesterase (Ace)

Baker and DeSalle (1997) used this locus to examine phylogenetic relationships in the Hawaiian Drosophila. We used it to look at recent divergences within the Drosophila mayaguana subcluster of the repleta species group O'Grady et al. 2002.

under construction

Alcohol Dehydrogenase (Adh)

This is the standard primer for drosophilid systematics. We use primers designed to hybridize in the first and third exons and span the entire second exon, as well as the first and thrid intron of this gene. The introns are useful for closely related divergences but are difficult to align above the species group level. Papers using these primers inlcude: O'Grady et al. (1998), O'Grady (1999), O'Grady and Kidwell 2002, Remsen and O'Grady (2002), and Zilversmit et al. (2002).

4682: 5' ACATYCAGCCAIGAGTTGAAYTTGTG 3'
4683: 5' CTGGGIGGCATTGGIYTSGACACCAC 3'

Amylase (Amy) 

T3/amyF1 5' ATTAACCCTCACTAAAGCGCCCCTGGTGGGARMGNTA 3'
T7/amyR1 5' AATACGACTCACTATAGCGCGCAGGCCCACNARYTCRCA 3'

bcDNA 

Zilversmit et al. (2002)

under construction

extra sex combs (esc) 

Zilversmit et al. (2002)

escL 5' GGCCATCAACGAGCTGAARTTYCAYCC 3'
escR 5' TTCCAGCACACGATGGCRTTYTCRCA 3'
T3/escL 5' ATTAACCCTCACTAAAGGGCCATCAACGACGTGAARTTYCAYCC 3'
T7/escR 5' AATACGACTCACTATAGCGAACCACTGCACGCAGTCNACRTARTT 3'

forkhead (fkh) 

Zilversmit et al. (2002)

T3/fkhL 5' ATTAACCCTCACTAAAGTCCCTACTCCTACATCTCCCTGATHACNATG 3'
T7/fkhR 5' AATACGACTCACTATAGCGCAGGTAGCAGCCGTTYTCRAACATRT 3'

glass (gl) 

This sequence was useful in inferring the phylogeny of the haleakalae species group of Hawaiian Drosophila (O'Grady and Zilversmit 2004) and among genera in Zilversmit et al. (2002).

glass1 5' TTTCGATTGCGGCGGNTGYTTYGA 3'
glass2 5'GCCGTGGTGCATGGTCATRTTCAT 3'
T3/glass1 5' ATTAACCCTCACTAAAGTTTCGATTGCGGCGGNTGYTTYGA 3'
T7/glass2 5' AATACGACTCACTATAGGCCGTGGTGCATGGTCATRTTCAT 3'

Glyceraldehyde phosphate dehydrogenase (Gpdh) 

Francisco Ayala's group has used this gene to estimate relationships within the Drosophilidae. O'Grady (1999) used this gene at the species group level. Remsen and O'Grady (2002) and Zilversmit et al. (2002) examined genus-level relationships in Drosophilidae using this primer pair.

under construction

HOM-C Genes 

DeSalle et al (2003) used degenerate primers to amplify several HOM-C homeobox genes from a number of drosophilid taxa. PCR products were cloned and shotgun sequenced to obtain sequences from the following loci: abdominal A, Antennapedia, sex combs reduced, ultrabithorax, deformed, fushi tarazu, abdominal B, labial and proboscopedia. These genes are all highly conserved and give good phylogenetic results at taxonomic levels above genus. 

HOM5L = 5' caracnytngarytngaraa 3'
HOM3R = 5' rttytcyttyttccayttcat 3'

hunchback (hb) 

This developmental gene was used by Baker and DeSalle (1997) to examine phylogenetic relationships in Hawaiian Drosophila. We used it to look at recent divergences within the Drosophila mayaguana subcluster of the repleta species group O'Grady et al. 2002.

under construction

Internal Transcribed Spacer (ITS1) 

ITS-1 is a transcribed spacer that sits between 28S and 5.8S in the ribosomal array. It was shown to be useful for rapid divergences in insects by Vogler and DeSalle (1994). We have used the sequence in two papers(O'Grady et al. 1998; O'Grady and Zilversmit 2004,) but don't generally use it because of difficulties aligning it with outgroup taxa.

under construction

kuzbanian (kuz)

Zilversmit et al. (2002)

under construction

lethal (2) neighbor of tid (l(2)not) 

Primer sequences for this locus have been published using the gene name ntid ( Bonacum et al. 2001). This sequence was useful in inferring the phylogeny of the haleakalae species group of Hawaiian Drosophila (O'Grady and Zilversmit 2004), as well as among genera in Zilversmit et al. (2002). Care should be taken in amplifying and aligning these sequences as this gene is a member of a multigene family and may have paralogy issues. 

ntidF1 5' GGGCCGCATCTTCGARCAYAARTGG 3'
ntidR1 5' TGGAGGGGTAGGTGTTCCARCARTA 3'
T3/ntidF1 5' ATTAACCCTCACTAAAGGGGCCGCATCTTCGARCAYAARTGG 3'
T7/ntidR1 5' AATACGACTCACTATAGTGGAGGGGTAGGTGTTCCARCARTA 3'

mago nashi (mago)

Zilversmit et al. (2002)

T3/magoL 5' ATTAACCCTCACTAAAGCCACAAGGGCAAGTTCGGNCAYGARTT 3'
T7/magoR 5' AATACGACTCACTATAGCACTTCAGGTCCTGCACCARRTARTARAA 3'

mastermind (mast) 

This gene was suggested by Brower and DeSalle (1994). We used it to look at recent divergences within the Drosophila mayaguana subcluster of the repleta species group O'Grady et al. 2002.

under construction

Mitochondrial assembly regulatory factor (Marf)

Primer sequences for this locus and a genus-level analysis have been published using the gene name CG3869 (Bonacum et al. 2001; Zilversmit et al. 2002). Oliveira et al. (2003) used the large non coding region of this locus to examine species level relationships within the Drosophila mayaguana cluster of the repleta species group. O'Grady and Zilversmit 2004 used this sequence to infer the phylogeny of the haleakalae species group of Hawaiian Drosophila. 

CG3869F 5' CCCAACATCTTCATCCTGAACAAYMGNTGGGA 3' 
CG3869R 5' GCGGACTGGGAGATGCAYTCYTCRAA 3'

POU domain protein 2 (pdm2) 

Zilversmit et al. (2002)

under construction

PSAP 

Zilversmit et al. (2002)

under construction

Rpt4 (Rpt4)

O'Grady and Zilversmit 2004 used this sequence to infer the phylogeny of the haleakalae species group of Hawaiian Drosophila.

under construction

sans fille (snf)

This set of primers amplifies both coding and non coding regions. Zilversmit et al. (2002) examined the relationships among genera in the Drosophilidae using the exon sequences and Oliveira et al. (2003) used the non coding portions of this locus to examine species level relationships within the Drosophila mayaguana cluster of the repleta species group. O'Grady and Zilversmit 2004 used this sequence to infer the phylogeny of the haleakalae species group of Hawaiian Drosophila. 

snfL 5' GAAGATGCGGGGCCARGCNTTYGT 3'
snfR 5' GAACAGCATGGACAGCATCATYTCRTT 3'
T3/snfL 5' ATTAACCCTCACTAAAGGAAGATGCGGGGCCARGCNTTYGT 3'
T7/snfR 5' AATACGACTCACTATAGGAACAGCATGGACAGCATCATYTCRTT 3'

seven in absentia (sina)

This sequence was useful in inferring the phylogeny of the haleakalae species group of Hawaiian Drosophila (O'Grady and Zilversmit 2004) and among genera in Zilversmit et al. (2002).

sia1 5' TCGAGTGCCCCGTGTGYTTYGAYTA 3'
sia2 5' GAAGTGGAAGCCGAAGCAGSWYTGCATCAT 3'
T3/sia1 5' ATTAACCCTCACTAAAGTCGAGTGCCCCGTGTGYTTYGAYTA 3'
T7/sia2 5' AATACGACTCACTATAGGAAGTGGAAGCCGAAGCAGSWYTGCATCAT 3'

Superoxide dismutase (Sod) 

Zilversmit et al. (2002)

under construction

vestigial (vg) 

This gene was suggested by Brower and DeSalle (1994). We used it to look at recent divergences within the Drosophila mayaguana subcluster of the repleta species group O'Grady et al. 2002.

under construction

wee 

Zilversmit et al. (2002)

weeL 5' GCCTGGGCCGAGGAYGAYCAYATG 3'
weeR 5' TCACGTGGCCCAGGTCNCCDATYTT 3'

wingless (wg)

Zilversmit et al. (2002)

wgL 5' GCAGTTCCGGAACCGGMGNTGGAAYTG 3'
wgR 5' GGACATGCCGTGGCACTTRCAYTCYTG 3'

Xanthene dehydrogenase (Xdh)

Zilversmit et al. (2002)

under construction