Chicken bc1 Sequencing efforts in Berkeley- LBNL and Pangene.

At the time the chicken structure was solved, no sequence information
was available for the chicken bc1 complex except that of cytochrome b.
(Cytochrome b is a favorite tool of molecular evolutionists, due to its
mitochondrial inheritance and hence lack of crossing-over)

In 1997 An-chian Chen and Sylvia Lo began the sequencing 
efforts, under the supervision of Hisao Yokota of LBNL.
They worked on the two remaining redox-factor-bearing
 subunits, cytochrome c1 and the Rieske Iron-sulfur protein. 

Despite extensive screening of hybridization conditions and
use of a number of primer pairs, Sylvia found nothing with
a sequence resembling cyt c1. In retrospect it looks like she 
had a tough project, as cyt c1 seems to be the least cooperative 
of the subunits (It could not be found in Goldman's cDNA library 
even given the correct sequence, and the chicken genome projects
- see below - only got two partial sequences (which overlap, 
fortunately)

An had better luck with the ISP. Her early results are described in 
this report, and yielded sequence for  residues 75-157 of the mature 
protein.  Later extended to 68-157 before she graduated and left.

Later Kyle Kwok and Amy Fang took over from An and Sylvia,
and Stan Goldman joined the group. Stan prepared size-fractionated
cDNA libraries from fresh chicken tissue, and began screening the libraries 
with primers designed from homologous sequences and with the ISP sequence 
obtained  by An.  The latter was successful, bringing up five positive clones, 
two of which were characterized. The first (bc41)  began at residue 88 and gave
accurate sequence to the stop codon and beyond. The second (bc45) included
the entire reading frame and thus the sequence for subunit 9 (ISP presequence)
as well as the mature ISP itself. 

Aligning the two reads of bc45 and one of bc41 with An's sequence gave 
us a sequence in which most of the residues were determined at least twice. 

Meanwhile two chicken genomics centers were producing large amounts of
DNA sequence and making it available to the public through BLAST servers
at their web site:
The Roslin institute chicken genome project
The University of Delaware Chick EST Database

Searching these databases initially gave sequences for everything except 
subunit II ("Core II") and cytochrome c1. There were a couple of short 
sequences from core protein II. 

I contracted with Pangene to fish out these two genes from 
Stan Goldman's library.  A scientist at Pangene discovered a partial cyt. c1
sequence from the Univ. Delaware in genbank, so they had exact sequences
to start with. Core protein II was straightforward and they provided the
entire sequence. However nothing matching cytochrome c1 could be found
in the library. Fortunately by now a second stretch of cyt c1 sequence has 
appeared from the genomics projects, and we have the entire sequence although
from a single read in most parts.

We have made the compiled sequences available, with links to the 
original sequences used in the compilation. We express our gratitude 
to the students and scientists who helped make this possible, and to 
Prof. Sung-Hou Kim for encouragement and financial support of the project.