TAICHUNG, TAIWAN
Inst. Molecular Biology, National Chung Hsing University

Map position of the centromere on chromosomes 1 and 9
--Lin, B-Y, Chang, S-J

B-A translocations with the most proximal breakpoints on both arms of chromosomes 1 and 9 were used to map the centromeric position on RFLP maps. The mapping strategy is based on the fact that the centromere is located between the breakpoints of the two most proximal translocations: one in the short arm and the other in the long arm of the same chromosome. The RFLP marker closest to but distal to the translocation breakpoint on both arms defines the map region of the centromere.

The marker position in relation to translocation breakpoints is determined by the terminal deficiencies generated by B-A translocations. Like the B chromosome, one of the two B-A translocation chromosomes, termed B-A, undergoes nondisjunction at the second pollen mitosis. This process generates two sperm: one with two B-As (hyperploid) and the other without any B-A (hypoploid). Upon fertilization with an egg carrying the normal chromosome complement, the latter sperm results in a hypoploid embryo deficient for the paternal copy of the chromosome arm distal to the translocation breakpoint. In other words, the hypoploid embryo associated with the terminally deficient chromosome arm is employed to generate DNA for RFLP analysis. For simplification of the RFLP analysis, both parents of the hypoploid progeny are in two different inbred backgrounds: the maternal parent is B73 and the paternal parent is L289. The RFLP analysis is done by Southern hybridization of the hypoploid DNA with each marker whose position in reference to the breakpoint is determined by the presence (proximal) or absence (distal) of the paternal signal.

Table 1 gives the mapping result of 10 RFLP markers on chromosome 1 by hypoploids of TB-1Sb and TB-1La. Three markers (asg45, csu3, and umc167) exhibit no paternal signal on the hypoploid of TB-1Sb, and six others (umc177a, bnl5.59, umc119, umc58, asg62, and bnl6.32) give no paternal signal on the hypoploid of TB- 1La. The paternal signal of one marker (umc67) is absent on the hypoploid of TB-1Sb and TB-1La, and none of the 10 markers have the paternal signal on the hypoploids of both translocations, indicating that they are not located in the region between the breakpoints of the two translocations, including the centromere. Disregarding umc67, current data map the centromere to the umc167-umc177a region, an interval of about 14 map units according to the map of Davis et al. (MNL 70:123-132, 1996). On the other hand, the fact that Burr et al. (MNL 69:247-256, 1995) mapped umc67 to the short arm of chromosome 1, places the centromere in the umc67-umc177a region that spans less than one map unit on the map of Davis et al. (1996). On the other hand, since the 1-B chromosome of TB-1Sb is associated with a rearrangement, and umc67, located near the end of the rearranged region, is deleted by multiple breakages during the formation of the translocation (Lin and Chang, MNL, this volume). In other words, umc67 is located on the long arm of chromosome 1, most proximal to the breakpoint of TB-1Sb. As a consequence, the centromere is located between umc167 and umc67, a region of 13 map units.

Table 1. Mapping 10 RFLP markers on chromosome 1 by hypoploids of TB-1Sb and TB-1La. (+, presence and -, absence of the paternal signal.)
 
RFLP markers Hypoploids  
  TB-1Sb TB-1La
asg45 - +
csu3(gfu) - +
umc167 - +
umc67 - -
umc177a (+)* -
bnl5.59 + -
umc119 + -
umc58 + -
asg62 + -
bnl6.32 + -
*presence of the non-L289 paternal signal (see following article).

Table 2 shows the results of mapping 12 RFLP markers by hypoploids of TB-9Sd and TB-9Lc. The paternal signal of five markers (umc109, umc113a, bnl3.06, umc247, and umc81) is absent on the hypoploid DNA of TB-9Sd, and that of six others (umc20, bnl5.04, umc114, bnl8.17, umc95, and csu54b) is not present on the hypoploid of TB-9Lc. The marker (bnl5.10) has no paternal signal on the hypoploid of TB-9Sd and TB-9Lc, and none of the markers have the paternal signal on the hypoploids of both translocations, implying all 12 markers are distal to the breakpoints of the two translocations. Without consideration of bnl5.10, current data place the centromere in the umc81-umc20 region, the length of which is about 2.5 map units in the Davis et al. map (1996) or 4 map units in the Burr et al. map (1995). Since bnl5.10 is located at the proximal end of a rearrangement on the 9-B chromosome of TB-9Lc, it was deleted by multiple breakages which occurred during the formation of the translocation (Lin and Chang, MNL, this volume). Accordingly, bnl5.10 is proximal to umc81, and the centromere is located in the bnl5.10-umc20 interval whose length can not be estimated because of the uncertain map position of bnl5.10. Burr et al. (1995) placed bnl5.10 on the short arm of chromosome 9, and Davis et al. (1996) mapped it to a position distal to umc20, which is on the long arm according to the data of this study.

Table 2. RFLP analysis of 12 markers on chromosome 9 by hypoploids of TB-9Sd and TB-9Lc. (+, presence and -, absence of the paternal signal.)
 
RFLP markers Hypoploids  
  TB-9Sd TB-9Lc
umc109 - +
umc113a - +
bnl3.06 - (+)*
umc247 - (+)*
umc81 - (+)*
bnl5.10 - -
umc20 + -
bnl5.04 (+)* -
umc114 (+)* -
bnl8.17 (+)* -
umc95 (+)* -
csu54b + -
*presence of the non-L289 paternal signal.


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