Table 2b.
CENPs assembly and centromere function on introduced alphoid DNAs: ectopic alphoid DNA integration sites
| Input DNA size (vector) | Cell lines | Multiplicity of input DNA | CENPs assembly (%)a | TSA | ||||||
|---|---|---|---|---|---|---|---|---|---|---|
| Left arm (copies) | Right arm (copies) | Total size of alphoid (kb)b | A | B | C | E | CENP-A assemblya (%) | Extra chromosome formationc (%) | ||
| Linear constructs | ||||||||||
| 70 kb (α7C5htel) | HT1-19d | 34 | 35 | 2400 | 17 | 20 | 17 | nt | 65 | 30 |
| HT2-1d | nt | nt | − | 13 | 39 | 23 | nt | 50 | 16 | |
| 50 kb (del.24) | del.24HT5–11 | 26 | 24 | 2500 | 13 | 100 | nt | nt | 45 | 20 |
| del.24HT5–20 | 27 | 32 | 1500 | 13 | 100 | nt | nt | 40 | 15 | |
| 30 kb (del.20) | del.20HT5–6 | 58 | 53 | 1700 | 25 | 100 | nt | 23 | nt | nt |
| del.20HT5–24 | 18 | 14 | 480 | 20 | 100 | nt | 25 | 40 | 20 | |
| 10 kb (del.22) | del.22HT1–3 | 94 | 101 | 980 | 0 | 1008 | 0 | nt | 23 | 0 |
| del.22HT5–7 | 53 | 38 | 480 | 0 | 100 | nt | nt | nt | nt | |
| del.22HT5–18 | 27 | 23 | 250 | 0 | 100 | nt | nt | 15 | 0 | |
| 9.4 kb (pNeo11.5) | W11.5HT1–16 | nt | 173 | 1600 | 0 | 100 | nt | nt | 10 | 0 |
| 9.4 kb (pNeo11.5htel) | W11.5htelHT1–23 | nt | 120 | 1100 | 0 | 100 | nt | nt | 10 | 0 |
| Circular constructs | ||||||||||
| 60 kb (pWTR11.32) | W0210R-1e | − | 14f | 1000 | 23 | 100 | nt | nt | nt | nt |
| 240 kb (pWTR11.128) | W11.128HT1–5 | − | 5f | 1200 | + | + | nt | nt | nt | nt |
| 240 kb (pWTR11.128) | W11.128HT4–3 | − | 6f | 1100 | + | + | nt | nt | nt | nt |
| 60 kb (pCBB11.1.32) | BR11.32HT2–25 | − | 42f | 4000 | + | + | nt | nt | nt | nt |
| 60 kb (pMTR11.32) | M1319e | − | 12f | 1100 | 0 | 0 | 0 | 0 | nt | nt |
| aPercentage of centromere protein assembly was assessed based on colocalized signals from indirect immunostaining of CENP and FISH analysis of vector probes (n>20). | ||||||||||
| bTotal size of alphoid DNA in multimer of input DNA was calculated by multiplying alphoid DNA size of input DNA by copy number of average of left arm and right arm. | ||||||||||
| cFormation of extra-chromosomes with active centromere formed on ectopic integration of input DNA were examined by immunostaining of CENP and FISH analysis of vector probes. | ||||||||||
| dThese cell lines were produced by Ikeno et al (1998). | ||||||||||
| eThese cell lines were produced by Ohzeki et al (2002). | ||||||||||
| fThese constructs are circular, therefore Neo gene in BAC vector sequence was quantified by real time-PCR. | ||||||||||
| gAmong them, 17% could be detected by the mixture of antibodies against CENP-B. | ||||||||||
| nt, not tested; +, assembly detected only with ChIP assay. | ||||||||||