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NEWSLETTER #2, December 22, 1989


Dear consortium members,

We have recieved the signed agreements from most of the groups who indicated an interest to join this CF Genetic Analysis Consortium. The total number of member is now 40; letters of agreement are expected to return from probably 10 more. It is a large number and we hope our FAX machines will not breakdown (see item 7). A membership list is attached (Those who have not return the signed agreement are also listed, but they will not receive this mailing). Please correct any errors and advise us as soon as possible.

1. There are several reports on the identification of additonal mutations.

(A) The first is that the Toronto group has detected another 3 bp deletion ([[Delta]]I507). While the normal DNA sequence there is AA AAT ATC ATC TTT GG, the mutant sequence is AA ATA ATC TTT GG. The mutation was first obtained by PCR cloning and sequencing, and confirmed by direct sequencing of the PCR product. The PCR primers for exon 10 have been described in the last newsletter (November 29, 1989). While we were ready to report to the consortium in early December, the Manchester group (McIntosh and Super) reported to have found the same (see letter attached). This mutation may be rather rare in the population (<1%) because each group has identified only one so far. In fact, the [[Delta]]I507 chromosome from Toronto belongs to the group III haplotype (the one that was thought to be a recurrent mutation) and the one from Manchester appears to be similar. If one compares [[Delta]]F508 and [[Delta]]I507, there is only a A to T change, thus explain the difficulty in distinguishing the two by hybridization, unless high stringency is used (2xSSC at 45[[ring]]). Surprisingly, the two could be easily distinguished by polyacrylamide gel electrophoresis- the corresponding heteroduplexes migrate very differently.

(B) Michael Dean reported a 2 bp insertion in exon 13 (see attached letter). The mutation (which he called CFins2566) may be detected by PCR and DdeI digestion. It is apparently also a rare one. No one else has reported this finding.

(C) Art Beaudet revealed the two normal chromosome variants that have amino acid changes at Ile506 and Phe508 (see letter). He reported his preliminary findings in Tampa. Michel Goossens found one of the changes more recently (see letter).

All these findings have been submitted or are being prepared for publication. Please regard these information as privileged communication until the papers are in print (ie. please do not discuss these data in public before seeing them in the journals), as we agreed according to the guidelines.

2. There are many reports on [[Delta]]F508 screening. Since each one was in a different format, I have summarized the data into a single table (instead of copying individual data sheets) (see attached Table). A number of European labs have decided to publish short reports in Human Genetics (see letter from Romeo); all European labs are invited to join. Again, please treat these population data as privileged communication.

3. The intron/exon boundary sequences for exon 14a and 15 are attached. We have distributed the sequences for exons 10, 11, 20 and 21 in the last newsletter to many members who returned their signed agreement earlier. Although my secretary has tried her best to send out the data to others who responded later, we might have missed a few. Please let us know if you have not received them.

4. Many investigators reported haplotype data with XV-2C and KM-19. Different degrees of association with [[Delta]]F508 have been observed, although the majority of [[Delta]]F508 goes with 12 (commonly known as the B haplotype). In our analysis with extended haplotypes, [[Delta]]F508 associates exclusively with group I which is defined mainly by DNA markers closely linked to the CF gene; 35% of the haplotype B CF chromosomes actually correspond to haplotypes groups other than Ia and 38% of the haplotype D belongs to group Ia, suggesting recombination between the gene and KM-19/XV-2C. Thus, it is not surprising to see [[Delta]]F508 on a D chromosome or other non-B chromosomes.

5. It has been suggested at the North American CF Conference in Tampa that we should try to find a most informative set of markers for future haplotyping purpose. Visual inspection of our data revealed that the sites J44(XbaI) and 10-1X.6(AccI or HaeIII) will distinguish I from II, and from the rest; T6/20(MspI) will distinguish III and IV from V; H1.3(NcoI) III from IV. All the probes have been deposited at ATCC and should be available for general distribution very soon. The J44(XbaI) polymorphism may be detected by PCR followed by restriction enzyme digestion. The two primers are 5'- CAA TGT GAT TGG TGA AAC TA and 5'- CTT CTC CTC CTA GAC ACC TGC AT.

6. Besides the typographical errors in the primer sequence and amino acid translations reported in the Science papers, we recently found another error in the nucleotide sequence- the nucleotide at position 1990 should be C, not A, and the corresponding amino acid should be His, not Asn. We apologize for these mistakes. The Toronto group also found 2 sequence polymorphisms. One is at nucleotide position 1540- the A residue can be substituted by G, changing the corresponding amino acid from Met to Val. The other is at position 2694- the T residue can be a G; although it does not change the amino acid the polymorphism may be detected by restriction enzymes AvaII or Sau96I. These changes are present in the normal population and show (absolute?) correlation with haplotypes.

7. Since our major communication means is via facsimile, please notify us immediately if there is a problem with your machine. This is particularly important because we are using an automatic dialing system; any non-responsive machine will cause a problem. It remains to be a problem with groups without FAX, but we have been using courier services.

8. I have received letters from some members complaining why nothing seemed to be happening after the signed consortium agreements were returned. I must admit that it was partly my fault because I had been overwhelmed by many other things during this period. One must, however, also understand that responses were only dripping in; we received two yesterday and one today. Many did not return the information sheet; some gave a long description of their overall interest instead. When you send in data, please be brief. Also, please indicate whether or not the information is for the consortium. If you fail to do so in the future, I will assume that it is for my information only. As before, I will hold off results on new mutations for 10 days before anouncement.

Wish you all have a Nice Holiday Season and a Happy New Year!


Reports on [[Delta]]F508 screening:

Source with [[Delta]]F508 without [[Delta]]F508 Haplotype Geographic/ethnic group

Beaudet 1 20 11(XV-KM) Houston, non-Jewish

314 57 12 non-hispanic

13 21

11 9 22

7 7 ?

9 23 12 Ashkenazic- Houston

1 non-12 Ashkenazic

10 1 12 Hispanic- Houston

1 2 non-12 Hispanic

1 2 ? Hispanic

Boué 136 64 not shown Paris

Cao 24 12(XV-KM) Sardinia

2 11

12 12

3 21

1 22

Cutting 16 27 not shown Baltimore- Am. Black

120 35 Baltimore- Caucasian

2 2 Baltimore- Oriental

Ferec 8 11(XV-KM) Brest

123 50 12

9 5 22

Goossens 14 11(XV-KM) Creteil

116 15 12

3 21

14 4 22

Halley (*) 3 6 11(XV-KM) Rotterdam

71 6 12

5 21

1 3 22

4 ?

12 10 ? Polish

Kitzis 5 8 11(XV-KM) Paris

104 17 12

15 21

4 11 22

Source with [[Delta]]F508 without [[Delta]]F508 Haplotype Geographic/ethnic group

McIntosh 1 5 11(XV-KM) Edinburgh

97 17 12

4 21

4 1 22

Naylor 6 8 Hispanic- San Antonio

10 5 Other Caucasian

Sheffer 1 11(XV-KM) Groningen

109 17 12

14 21

3 4 22

Schwartz 249 43 Copenhagen

Tsui 313 129 not done Toronto- heterogeneous

for new ones Caucasian

Tümmler 40 11121 (J311/M-KM-XV-metH/T-metD/T)

10 12121 Hannover

10 1 11122

10 12122

3 21121

4 21122

1 1 11222

2 11111

1 11112

1 12111

1 22121

1 11212

1 12212

1 21211

1 12221

1 21221

* from poster in ASHG, Baltimore